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Find video protocols related to scientific articles indexed in Pubmed.
Reversible H atom abstraction catalyzed by the radical S-adenosylmethionine enzyme HydG.
J. Am. Chem. Soc.
PUBLISHED: 09-10-2014
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The organometallic H-cluster at the active site of [FeFe]-hydrogenases is synthesized by three accessory proteins, two of which are radical S-adenosylmethionine enzymes (HydE, HydG) and one of which is a GTPase (HydF). In this work we probed the specific role of H atom abstraction in HydG-catalyzed carbon monoxide and cyanide production from tyrosine. The isotope distributions of 5'-deoxyadenosine and p-cresol were evaluated using deuterium-labeled tyrosine substrates in H2O and D2O. The observation of multiply deuterated 5'-deoxyadenosine and deuterated S-adenosylmethionine when the reaction is carried out in D2O provides evidence for a 5'-deoxyadenosyl radical-mediated abstraction of a hydrogen atom from a solvent-exchangeable position as a reversible event.
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[FeFe]-hydrogenase maturation.
Biochemistry
PUBLISHED: 06-16-2014
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Hydrogenases are metalloenzymes that catalyze the reversible reduction of protons at unusual metal centers. This Current Topic discusses recent advances in elucidating the steps involved in the biosynthesis of the complex metal cluster at the [FeFe]-hydrogenase (HydA) active site, known as the H-cluster. The H-cluster is composed of a 2Fe subcluster that is anchored within the active site by a bridging cysteine thiolate to a [4Fe-4S] cubane. The 2Fe subcluster contains carbon monoxide, cyanide, and bridging dithiolate ligands. H-cluster biosynthesis is now understood to occur stepwise; standard iron-sulfur cluster assembly machinery builds the [4Fe-4S] cubane of the H-cluster, while three specific maturase enzymes known as HydE, HydF, and HydG assemble the 2Fe subcluster. HydE and HydG are both radical S-adenosylmethionine enzymes that interact with an iron-sulfur cluster binding GTPase scaffold, HydF, during the construction of the 2Fe subcluster moiety. In an unprecedented biochemical reaction, HydG cleaves tyrosine and decomposes the resulting dehydroglycine into carbon monoxide and cyanide ligands. The role of HydE in the biosynthetic pathway remains undefined, although it is hypothesized to be critical for the synthesis of the bridging dithiolate. HydF is the site where the complete 2Fe subcluster is formed and ultimately delivered to the immature hydrogenase protein in the final step of [FeFe]-hydrogenase maturation. This work addresses the roles of and interactions among HydE, HydF, HydG, and HydA in the formation of the mature [FeFe]-hydrogenase.
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Solution phase dynamics of the DNA repair enzyme spore photoproduct lyase as probed by H/D exchange.
FEBS Lett.
PUBLISHED: 05-05-2014
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Spore photoproduct lyase (SPL) catalyzes the repair of the UV lesion spore photoproduct (SP) in a reaction dependent on S-adenosyl-L-methionine (SAM). We have utilized H/D exchange to show that in the presence of SAM, a significant reduction in H/D exchange is observed upon binding SPTpT or undamaged oligonucleotide, indicating a shift of 20 or 10 amide protons, respectively, from a rapidly-exchangable state to a fully-protected conformation. In the absence of SAM, neither the oligonucleotide nor the SPTpT produce a significant perturbation in H/D exchange, indicating SAM is a requisite binding partner. Performing the same experiments in aerobic conditions reduced the magnitude of ligand-induced structural changes, consistent with the importance of the oxygen-sensitive iron-sulfur cluster for SAM and substrate binding.
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Nurse practitioners can effectively deliver pain coping skills training to osteoarthritis patients with chronic pain: A randomized, controlled trial.
Pain
PUBLISHED: 03-18-2014
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A multisite, randomized, controlled clinical effectiveness trial was conducted for osteoarthritis patients with chronic pain of the knee or hip. Adult health nurse practitioners provided a 10-session intervention, pain coping skills training (PCST), in patients' doctors' offices (N=129 patients); the control group received usual care (N=127 patients). Primary outcomes assessed at baseline, posttreatment, 6-month follow-up, and 12-month follow-up were: pain intensity, physical functioning, psychological distress, self-efficacy, catastrophizing, use of coping strategies, and quality of life. Secondary measures included fatigue, social functioning, health satisfaction, and use of pain medication. Methods favoring external validity, consistent with pragmatic, effectiveness research, were utilized. Primary ITT and secondary per-protocol analyses were conducted. Attrition was within the expected range: 11% at posttreatment and 29% at 12-month follow-up; rates did not differ between groups. Omnibus ITT analyses across all assessment points indicated significant improvement for the PCST group compared with the control group for pain intensity, physical functioning, psychological distress, use of pain coping strategies, and self-efficacy, as well as fatigue, satisfaction with health, and reduced use of pain medication. Treatment effects were robust to covariates (demographics and clinical sites). Trends in the outcomes across the assessments were examined. All outcomes, except for self-efficacy, were maintained through the 12-month follow-up; effects for self-efficacy degraded over time. Per-protocol analyses did not yield greater effect sizes. Comparisons of PCST patients who were more vs less treatment adherent suggested greater effectiveness for patients with high adherence. Results support the effectiveness of nurse practitioner delivery of PCST for chronic osteoarthritis pain.
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H-Cluster assembly during maturation of the [FeFe]-hydrogenase.
J. Biol. Inorg. Chem.
PUBLISHED: 02-19-2014
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The organometallic H-cluster at the active site of the [FeFe]-hydrogenase serves as the site of reversible binding and reduction of protons to produce H2. The H-cluster is unique in biology, and consists of a 2Fe subcluster tethered to a typical [4Fe-4S] cluster by a single cysteine ligand. The remaining ligands to the 2Fe subcluster include three carbon monoxides, two cyanides, and a dithiomethylamine. This mini-review will focus on the significant advances in recent years in understanding the pathway for H-cluster biosynthesis, as well as the structures, roles, and mechanisms of the three enzymes directly involved.
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Ecological validity and clinical utility of Patient-Reported Outcomes Measurement Information System (PROMIS®) instruments for detecting premenstrual symptoms of depression, anger, and fatigue.
J Psychosom Res
PUBLISHED: 01-28-2014
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This study examined the ecological validity and clinical utility of NIH Patient Reported-Outcomes Measurement Information System (PROMIS®) instruments for anger, depression, and fatigue in women with premenstrual symptoms.
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Glycyl radical activating enzymes: structure, mechanism, and substrate interactions.
Arch. Biochem. Biophys.
PUBLISHED: 01-20-2014
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The glycyl radical enzyme activating enzymes (GRE-AEs) are a group of enzymes that belong to the radical S-adenosylmethionine (SAM) superfamily and utilize a [4Fe-4S] cluster and SAM to catalyze H-atom abstraction from their substrate proteins. GRE-AEs activate homodimeric proteins known as glycyl radical enzymes (GREs) through the production of a glycyl radical. After activation, these GREs catalyze diverse reactions through the production of their own substrate radicals. The GRE-AE pyruvate formate lyase activating enzyme (PFL-AE) is extensively characterized and has provided insights into the active site structure of radical SAM enzymes including GRE-AEs, illustrating the nature of the interactions with their corresponding substrate GREs and external electron donors. This review will highlight research on PFL-AE and will also discuss a few GREs and their respective activating enzymes.
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The association of somatic arousal with the symptoms of upper airway resistance syndrome.
Sleep Med.
PUBLISHED: 01-14-2014
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We tested the hypothesis that the symptoms of upper airway resistance syndrome (UARS) are manifestations of chronic stress. To accomplish this, we utilized the score on a self-report questionnaire for somatic arousal (a component of stress) to compare somatic arousal between UARS patients and healthy controls and, among all participants, to correlate the level of somatic arousal with the severity of UARS symptoms.
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Combined Mössbauer spectroscopic, multi-edge X-ray absorption spectroscopic, and density functional theoretical study of the radical SAM enzyme spore photoproduct lyase.
J. Biol. Inorg. Chem.
PUBLISHED: 01-13-2014
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Spore photoproduct lyase (SPL), a member of the radical S-adenosyl-L-methionine (SAM) superfamily, catalyzes the direct reversal of the spore photoproduct, a thymine dimer specific to bacterial spores, to two thymines. SPL requires SAM and a redox-active [4Fe-4S] cluster for catalysis. Mössbauer analysis of anaerobically purified SPL indicates the presence of a mixture of cluster states with the majority (40 %) as [2Fe-2S](2+) clusters and a smaller amount (15 %) as [4Fe-4S](2+) clusters. On reduction, the cluster content changes to primarily (60 %) [4Fe-4S](+). The speciation information from Mössbauer data allowed us to deconvolute iron and sulfur K-edge X-ray absorption spectra to uncover electronic (X-ray absorption near-edge structure, XANES) and geometric (extended X-ray absorption fine structure, EXAFS) structural features of the Fe-S clusters, and their interactions with SAM. The iron K-edge EXAFS data provide evidence for elongation of a [2Fe-2S] rhomb of the [4Fe-4S] cluster on binding SAM on the basis of an Fe···Fe scatterer at 3.0 Å. The XANES spectra of reduced SPL in the absence and presence of SAM overlay one another, indicating that SAM is not undergoing reductive cleavage. The X-ray absorption spectroscopy data for SPL samples and data for model complexes from the literature allowed the deconvolution of contributions from [2Fe-2S] and [4Fe-4S] clusters to the sulfur K-edge XANES spectra. The analysis of pre-edge features revealed electronic changes in the Fe-S clusters as a function of the presence of SAM. The spectroscopic findings were further corroborated by density functional theory calculations that provided insights into structural and electronic perturbations that can be correlated by considering the role of SAM as a catalyst or substrate.
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Pyruvate Formate-Lyase and its Activation by Pyruvate Formate-Lyase Activating Enzyme.
J. Biol. Chem.
PUBLISHED: 12-12-2013
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The activation of pyruvate formate-lyase (PFL) by pyruvate formate-lyase activating enzyme (PFL-AE) involves formation of a specific glycyl radical on PFL by the PFL-AE in a reaction requiring S-adenosylmethionine (SAM). Surface plasmon resonance (SPR) experiments were performed under anaerobic conditions on the oxygen sensitive PFL-AE to determine the kinetics and equilibrium constant for its interaction with PFL. These experiments show that the interaction is very slow and rate-limited by large conformational changes. A novel SAM binding assay was used to accurately determine the equilibrium constants for SAM binding to PFL-AE alone and in complex with PFL. The PFL-AE bound SAM with the same affinity (~ 6 ?M) regardless of the presence or absence of PFL. Activation of PFL in the presence of its substrate pyruvate or the analogue oxamate resulted in stoichiometric conversion of the [4Fe-4S]1+ cluster to the glycyl radical on PFL, however 3.7-fold less activation was achieved in the absence of these small molecules, demonstrating that pyruvate or oxamate are required for optimal activation. Finally, in vivo concentrations of the entire PFL system were calculated to estimate the amount of bound protein in the cell. PFL, PFL-AE, and SAM are essentially fully bound in vivo, whereas electron donor proteins are partially bound.
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Biochemical and Kinetic Characterization of Radical S-Adenosyl-l-methionine Enzyme HydG.
Biochemistry
PUBLISHED: 11-20-2013
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The radical S-adenosyl-l-methionine (AdoMet) enzyme HydG is one of three maturase enzymes involved in [FeFe]-hydrogenase H-cluster assembly. It catalyzes l-tyrosine cleavage to yield the H-cluster cyanide and carbon monoxide ligands as well as p-cresol. Clostridium acetobutylicum HydG contains the conserved CX3CX2C motif coordinating the AdoMet binding [4Fe-4S] cluster and a C-terminal CX2CX22C motif proposed to coordinate a second [4Fe-4S] cluster. To improve our understanding of the roles of each of these iron-sulfur clusters in catalysis, we have generated HydG variants lacking either the N- or C-terminal cluster and examined these using spectroscopic and kinetic methods. We have used iron analyses, UV-visible spectroscopy, and electron paramagnetic resonance (EPR) spectroscopy of an N-terminal C96/100/103A triple HydG mutant that cannot coordinate the radical AdoMet cluster to unambiguously show that the C-terminal cysteine motif coordinates an auxiliary [4Fe-4S] cluster. Spectroscopic comparison with a C-terminally truncated HydG (?CTD) harboring only the N-terminal cluster demonstrates that both clusters have similar UV-visible and EPR spectral properties, but that AdoMet binding and cleavage occur only at the N-terminal radical AdoMet cluster. To elucidate which steps in the catalytic cycle of HydG require the auxiliary [4Fe-4S] cluster, we compared the Michaelis-Menten constants for AdoMet and l-tyrosine for reconstituted wild-type, C386S, and ?CTD HydG and demonstrate that these C-terminal modifications do not affect the affinity for AdoMet but that the affinity for l-tyrosine is drastically reduced compared to that of wild-type HydG. Further detailed kinetic characterization of these HydG mutants demonstrates that the C-terminal cluster and residues are not essential for l-tyrosine cleavage to p-cresol but are necessary for conversion of a tyrosine-derived intermediate to cyanide and CO.
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Measuring daily fatigue using a brief scale adapted from the Patient-Reported Outcomes Measurement Information System (PROMIS(®)).
Qual Life Res
PUBLISHED: 10-01-2013
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Daily assessments can provide insight into the temporal characteristics of fatigue. They can demonstrate consistency or reveal variability, as when fatigue changes with the underlying medical condition, improves with therapy, or worsens as a medication side effect. We adapted a fatigue measure from the Patient-Reported Outcomes Measurement Information System (PROMIS(®)) for daily assessment and examined its psychometric properties in a month-long prospective study.
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Accuracy of 30-Day Recall for Components of Sexual Function and the Moderating Effects of Gender and Mood.
J Sex Med
PUBLISHED: 06-26-2013
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Despite the ubiquity of 1-month recall periods for measures of sexual function, there is limited evidence for how well recalled responses correspond to individuals actual daily experiences.
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EPR and FTIR analysis of the mechanism of H2 activation by [FeFe]-hydrogenase HydA1 from Chlamydomonas reinhardtii.
J. Am. Chem. Soc.
PUBLISHED: 04-24-2013
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While a general model of H2 activation has been proposed for [FeFe]-hydrogenases, the structural and biophysical properties of the intermediates of the H-cluster catalytic site have not yet been discretely defined. Electron paramagnetic resonance (EPR) spectroscopy and Fourier transform infrared (FTIR) spectroscopy were used to characterize the H-cluster catalytic site, a [4Fe-4S]H subcluster linked by a cysteine thiolate to an organometallic diiron subsite with CO, CN, and dithiolate ligands, in [FeFe]-hydrogenase HydA1 from Chlamydomonas reinhardtii (CrHydA1). Oxidized CrHydA1 displayed a rhombic 2.1 EPR signal (g = 2.100, 2.039, 1.997) and an FTIR spectrum previously assigned to the oxidized H-cluster (Hox). Reduction of the Hox sample with 100% H2 or sodium dithionite (NaDT) nearly eliminated the 2.1 signal, which coincided with appearance of a broad 2.3-2.07 signal (g = 2.3-2.07, 1.863) and/or a rhombic 2.08 signal (g = 2.077, 1.935, 1.880). Both signals displayed relaxation properties similar to those of [4Fe-4S] clusters and are consistent with an S = 1/2 H-cluster containing a [4Fe-4S]H(+) subcluster. These EPR signals were correlated with differences in the CO and CN ligand modes in the FTIR spectra of H2- and NaDT-reduced samples compared with Hox. The results indicate that reduction of [4Fe-4S]H from the 2+ state to the 1+ state occurs during both catalytic H2 activation and proton reduction and is accompanied by structural rearrangements of the diiron subsite CO/CN ligand field. Changes in the [4Fe-4S]H oxidation state occur in electron exchange with the diiron subsite during catalysis and mediate electron transfer with either external carriers or accessory FeS clusters.
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Flavodoxin cofactor binding induces structural changes that are required for protein-protein interactions with NADP(+) oxidoreductase and pyruvate formate-lyase activating enzyme.
Biochim. Biophys. Acta
PUBLISHED: 03-25-2013
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Flavodoxin (Fld) conformational changes, thermal stability, and cofactor binding were studied using circular dichroism (CD), isothermal titration calorimetry (ITC), and limited proteolysis. Thermodynamics of apo and holo-Fld folding were examined to discern the features of this important electron transfer protein and to provide data on apo-Fld. With the exception of fluorescence and UV-vis binding experiments with its cofactor flavin mononucleotide (FMN), apo-Fld is almost completely uncharacterized in Escherichia coli. Fld is more structured when the FMN cofactor is bound; the association is tight and driven by enthalpy of binding. Surface plasmon resonance binding experiments were carried out under anaerobic conditions for both apo- and holo-Fld and demonstrate the importance of structure and conformation for the interaction with binding partners. Holo-Fld is capable of associating with NADP(+)-dependent flavodoxin oxidoreductase (FNR) and pyruvate formate-lyase activating enzyme (PFL-AE) whereas there is no detectable interaction between apo-Fld and either protein. Limited proteolysis experiments were analyzed by LC-MS to identify the regions in Fld that are involved in conformation changes upon cofactor binding. Docking software was used to model the Fld/PFL-AE complex to understand the interactions between these two proteins and gain insight into electron transfer reactions from Fld to PFL-AE.
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Temporal trends in symptom experience predict the accuracy of recall PROs.
J Psychosom Res
PUBLISHED: 03-10-2013
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Patient-reported outcome measures with reporting periods of a week or more are often used to evaluate the change of symptoms over time, but the accuracy of recall in the context of change is not well understood. This study examined whether temporal trends in symptoms that occur during the reporting period impact the accuracy of 7-day recall reports.
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Snapshot From Superstorm Sandy: American Red Cross Mental Health Risk Surveillance in Lower New York State.
Ann Emerg Med
PUBLISHED: 02-19-2013
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Disasters often cause psychological injury, as well as dramatic physical damage. Epidemiologic research has identified a set of disaster experiences and predisposing characteristics that place survivors at risk for post traumatic stress disorder (PTSD), depression, and anxiety. Rapid triage of at-risk survivors could have benefits for individual and population-level outcomes. We examine American Red Cross mental health risk surveillance data collected from October 29 to November 20, 2012, immediately after Hurricane Sandy in 8 lower New York State counties to evaluate the feasibility and utility of collecting these data.
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A comparison of inspiratory airflow dynamics during sleep between upper airway resistance syndrome patients and healthy controls.
Sleep Breath
PUBLISHED: 01-23-2013
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A test of the hypothesis that upper airway resistance syndrome (UARS) patients have an increased prevalence of inspiratory airflow limitation (IFL) during sleep compared to healthy controls.
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Validity and reliability of patient-reported outcomes measurement information system instruments in osteoarthritis.
Arthritis Care Res (Hoboken)
PUBLISHED: 01-17-2013
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Evaluation of known-group validity, ecological validity, and test-retest reliability of 4 domain instruments from the Patient-Reported Outcomes Measurement Information System (PROMIS) in osteoarthritis (OA) patients.
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S K-edge XAS and DFT calculations on SAM dependent pyruvate formate-lyase activating enzyme: nature of interaction between the Fe4S4 cluster and SAM and its role in reactivity.
J. Am. Chem. Soc.
PUBLISHED: 10-28-2011
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S K-edge X-ray absorption spectroscopy on the resting oxidized and the S-adenosyl-l-methionine (SAM) bound forms of pyruvate formate-lyase activating enzyme are reported. The data show an increase in pre-edge intensity, which is due to additional contributions from sulfide and thiolate of the Fe(4)S(4) cluster into the C-S ?* orbital. This experimentally demonstrates that there is a backbonding interaction between the Fe(4)S(4) cluster and C-S ?* orbitals of SAM in this inner sphere complex. DFT calculations that reproduce the data indicate that this backbonding is enhanced in the reduced form and that this configurational interaction between the donor and acceptor orbitals facilitates the electron transfer from the cluster to the SAM, which otherwise has a large outer sphere electron transfer barrier. The energy of the reductive cleavage of the C-S bond is sensitive to the dielectric of the protein in the immediate vicinity of the site as a high dielectric stabilizes the more charge separated reactant increasing the reaction barrier. This may provide a mechanism for generation of the 5-deoxyadenosyl radical upon substrate binding.
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Experimental induction of psychogenic illness in the context of a medical event and media exposure.
Am J Disaster Med
PUBLISHED: 08-30-2011
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Mass psychogenic illness can be a significant problem for triage and hospital surge in disasters; however, research has been largely limited to posthoc observational reports. Reports on the impact of public media during a disaster have suggested both salutary as well as iatrogenic psychological effects. This study was designed to determine if psychogenic illness can be evoked and if media will exacerbate it in a plausible, controlled experiment among healthy community adults.
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Treatment expectation for pain coping skills training: relationship to osteoarthritis patients baseline psychosocial characteristics.
Clin J Pain
PUBLISHED: 08-05-2011
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This study examined predictors of treatment expectation among osteoarthritis (OA) patients in a multisite clinical trial of pain coping skills training (CST).
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Insights into [FeFe]-hydrogenase structure, mechanism, and maturation.
Structure
PUBLISHED: 04-01-2011
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Hydrogenases are metalloenzymes that are key to energy metabolism in a variety of microbial communities. Divided into three classes based on their metal content, the [Fe]-, [FeFe]-, and [NiFe]-hydrogenases are evolutionarily unrelated but share similar nonprotein ligand assemblies at their active site metal centers that are not observed elsewhere in biology. These nonprotein ligands are critical in tuning enzyme reactivity, and their synthesis and incorporation into the active site clusters require a number of specific maturation enzymes. The wealth of structural information on different classes and different states of hydrogenase enzymes, biosynthetic intermediates, and maturation enzymes has contributed significantly to understanding the biochemistry of hydrogen metabolism. This review highlights the unique structural features of hydrogenases and emphasizes the recent biochemical and structural work that has created a clearer picture of the [FeFe]-hydrogenase maturation pathway.
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Biosynthesis of complex iron-sulfur enzymes.
Curr Opin Chem Biol
PUBLISHED: 01-31-2011
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Recent advances in our understanding of the mechanisms for the biosynthesis of the complex iron-sulfur (Fe-S) containing prosthetic groups associated with [FeFe]-hydrogenases and nitrogenases have revealed interesting parallels. The biosynthesis of the H-cluster ([FeFe]-hydrogenase) and the FeMo-co (nitrogenase) occurs through a coordinated process that involves the modification of Fe-S cluster precursors synthesized by the general host cell machinery (Isc/Suf). Key modifications to the Fe-S precursors are introduced by the activity of radical S-adenosylmethionine (SAM) enzymes on unique scaffold proteins. The transfer of the modified clusters to a cofactor-less structural apo-protein completes maturation. Together these features provide the basis for establishing unifying paradigms for complex Fe-S cluster biosynthesis for these enzymes.
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[FeFe]-hydrogenase maturation: HydG-catalyzed synthesis of carbon monoxide.
J. Am. Chem. Soc.
PUBLISHED: 06-23-2010
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Biosynthesis of the unusual organometallic H-cluster at the active site of the [FeFe]-hydrogenase requires three accessory proteins, two of which are radical AdoMet enzymes (HydE, HydG) and one of which is a GTPase (HydF). We demonstrate here that HydG catalyzes the synthesis of CO using tyrosine as a substrate. CO production was detected by using deoxyhemoglobin as a reporter and monitoring the appearance of the characteristic visible spectroscopic features of carboxyhemoglobin. Assays utilizing (13)C-tyrosine were analyzed by FTIR to confirm the production of HbCO and to demonstrate that the CO product was synthesized from tyrosine. CO ligation is a common feature at the active sites of the [FeFe], [NiFe], and [Fe]-only hydrogenases; however, this is the first report of the enzymatic synthesis of CO in hydrogenase maturation.
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Pyruvate formate-lyase, evidence for an open conformation favored in the presence of its activating enzyme.
J. Biol. Chem.
PUBLISHED: 06-22-2010
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Pyruvate formate-lyase-activating enzyme (PFL-AE) activates pyruvate formate-lyase (PFL) by generating a catalytically essential radical on Gly-734 of PFL. Crystal structures of unactivated PFL reveal that Gly-734 is buried 8 A from the surface of the protein in what we refer to here as the closed conformation of PFL. We provide here the first experimental evidence for an alternate open conformation of PFL in which: (i) the glycyl radical is significantly less stable; (ii) the activated enzyme exhibits lower catalytic activity; (iii) the glycyl radical undergoes less H/D exchange with solvent; and (iv) the T(m) of the protein is decreased. The evidence suggests that in the open conformation of PFL, the Gly-734 residue is located not in its buried position in the enzyme active site but rather in a more solvent-exposed location. Further, we find that the presence of the PFL-AE increases the proportion of PFL in the open conformation; this observation supports the idea that PFL-AE accesses Gly-734 for direct hydrogen atom abstraction by binding to the Gly-734 loop in the open conformation, thereby shifting the closed <--> open equilibrium of PFL to the right. Together, our results lead to a model in which PFL can exist in either a closed conformation, with Gly-734 buried in the active site of PFL and harboring a stable glycyl radical, or an open conformation, with Gly-734 more solvent-exposed and accessible to the PFL-AE active site. The equilibrium between these two conformations of PFL is modulated by the interaction with PFL-AE.
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Synthesis of the 2Fe subcluster of the [FeFe]-hydrogenase H cluster on the HydF scaffold.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 05-24-2010
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The organometallic H cluster at the active site of [FeFe]-hydrogenase consists of a 2Fe subcluster coordinated by cyanide, carbon monoxide, and a nonprotein dithiolate bridged to a [4Fe-4S] cluster via a cysteinate ligand. Biosynthesis of this cluster requires three accessory proteins, two of which (HydE and HydG) are radical S-adenosylmethionine enzymes. The third, HydF, is a GTPase. We present here spectroscopic and kinetic studies of HydF that afford fundamental new insights into the mechanism of H-cluster assembly. Electron paramagnetic spectroscopy reveals that HydF binds both [4Fe-4S] and [2Fe-2S] clusters; however, when HydF is expressed in the presence of HydE and HydG (HydF(EG)), only the [4Fe-4S] cluster is observed by EPR. Insight into the fate of the [2Fe-2S] cluster harbored by HydF is provided by FTIR, which shows the presence of carbon monoxide and cyanide ligands in HydF(EG). The thorough kinetic characterization of the GTPase activity of HydF shows that activity can be gated by monovalent cations and further suggests that GTPase activity is associated with synthesis of the 2Fe subcluster precursor on HydF, rather than with transfer of the assembled precursor to hydrogenase. Interestingly, we show that whereas the GTPase activity is independent of the presence of the FeS clusters on HydF, GTP perturbs the EPR spectra of the clusters, suggesting communication between the GTP- and cluster-binding sites. Together, the results indicate that the 2Fe subcluster of the H cluster is synthesized on HydF from a [2Fe-2S] cluster framework in a process requiring HydE, HydG, and GTP.
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Interference with activities due to pain and fatigue: accuracy of ratings across different reporting periods.
Qual Life Res
PUBLISHED: 05-21-2010
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This study examined the impact of different reporting period lengths on the accuracy of items measuring interference due to pain and fatigue with work, walking, and relations with others.
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A snapshot of the age distribution of psychological well-being in the United States.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 05-17-2010
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Psychological well-being (WB) includes a persons overall appraisal of his or her life (Global WB) and affective state (Hedonic WB), and it is considered a key aspect of the health of individuals and groups. Several cross-sectional studies have documented a relation between Global WB and age. Little is known, however, about the age distribution of Hedonic WB. It may yield a different view of aging because it is less influenced by the cognitive reconstruction inherent in Global WB measures and because it includes both positive and negative components of WB. In this study we report on both Global and Hedonic WB assessed in a 2008 telephone survey of 340,847 people in the United States. Consistent with prior studies, Global WB and positive Hedonic WB generally had U-shaped age profiles showing increased WB after the age of 50 years. However, negative Hedonic WB variables showed distinctly different and stronger patterns: Stress and Anger steeply declined from the early 20s, Worry was elevated through middle age and then declined, and Sadness was essentially flat. Unlike a prior study, men and women had very similar age profiles of WB. Several measures that could plausibly covary with the age-WB association (e.g., having children at home) did not alter the age-WB patterns. Global and Hedonic WB measures appear to index different aspects of WB over the lifespan, and the postmidlife increase in WB, especially in Hedonic WB, deserves continued exploration.
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Single momentary assessments are not reliable outcomes for clinical trials.
Contemp Clin Trials
PUBLISHED: 05-03-2010
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Patient reported outcomes (PROs) play an essential role in clinical trials, though questions have been raised about the accuracy of PROs using long recall periods. This paper examines the utility of a PRO employing a single momentary assessment of pain in a sample of community rheumatology patients. We explore the accuracy and reliability of a single assessment versus the average of multiple assessments taken over 1-week, which is considered a common outcome reporting period. A secondary analysis of 128 patients who monitored their pain intensity with momentary data collections several times a day for a week and 3 months later for another week allowed a comparison of randomly-selected single momentary assessments with the average of many assessments from the week. Results from cross-sectional analyses of the first week were that levels of pain measured by single points were not significantly different than the week average in 4 of 5 analyses, but these single-point assessments had much higher variance. Correlations of single-point and week averages were below 0.70. Longitudinal analysis of change scores across 3 months also demonstrated considerable unreliability of single-point measures, thus the statistical power generated by single-point assessments was considerably less than the more reliable week average. Our conclusion is that single momentary assessments, at least for representing an outcome over a period of a week, are not ideal measures. We discuss alternative measurement strategies for efficiently collecting PRO data for a 1-week period using end-of-day diaries or 7-day recall measures.
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The effect of nasal continuous positive airway pressure on the symptoms of Gulf War illness.
Sleep Breath
PUBLISHED: 04-28-2010
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We performed a pilot study to determine whether nasal continuous positive airway pressure (CPAP) alleviates the symptoms of veterans with Gulf War illness (GWI) and sleep disordered breathing (SDB).
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Inspiratory airflow dynamics during sleep in veterans with Gulf War illness: a controlled study.
Sleep Breath
PUBLISHED: 04-28-2010
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To determine whether veterans with Gulf War Illness (GWI) are distinguished by sleep-disordered breathing, we compared inspiratory airflow dynamics during sleep between veterans with GWI and asymptomatic veterans of the first Gulf War.
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An efficient deprotection of N-trimethylsilylethoxymethyl (SEM) groups from dinucleosides and dinucleotides.
Nucleosides Nucleotides Nucleic Acids
PUBLISHED: 04-15-2010
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A convenient and efficient method for deprotection of N-(trimethyl)silylethoxymethyl (SEM) groups from thymidine dinucleoside and dinucleotide has been achieved. The SEM groups were easily removed in excellent yields from protected nucleosides, dinucleosides, and dinucleotides.
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Stepwise [FeFe]-hydrogenase H-cluster assembly revealed in the structure of HydA(DeltaEFG).
Nature
PUBLISHED: 03-05-2010
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Complex enzymes containing Fe-S clusters are ubiquitous in nature, where they are involved in a number of fundamental processes including carbon dioxide fixation, nitrogen fixation and hydrogen metabolism. Hydrogen metabolism is facilitated by the activity of three evolutionarily and structurally unrelated enzymes: the [NiFe]-hydrogenases, [FeFe]-hydrogenases and [Fe]-hydrogenases (Hmd). The catalytic core of the [FeFe]-hydrogenase (HydA), termed the H-cluster, exists as a [4Fe-4S] subcluster linked by a cysteine thiolate to a modified 2Fe subcluster with unique non-protein ligands. The 2Fe subcluster and non-protein ligands are synthesized by the hydrogenase maturation enzymes HydE, HydF and HydG; however, the mechanism, synthesis and means of insertion of H-cluster components remain unclear. Here we show the structure of HydA(DeltaEFG) (HydA expressed in a genetic background devoid of the active site H-cluster biosynthetic genes hydE, hydF and hydG) revealing the presence of a [4Fe-4S] cluster and an open pocket for the 2Fe subcluster. The structure indicates that H-cluster synthesis occurs in a stepwise manner, first with synthesis and insertion of the [4Fe-4S] subcluster by generalized host-cell machinery and then with synthesis and insertion of the 2Fe subcluster by specialized hydE-, hydF- and hydG-encoded maturation machinery. Insertion of the 2Fe subcluster presumably occurs through a cationically charged channel that collapses following incorporation, as a result of conformational changes in two conserved loop regions. The structure, together with phylogenetic analysis, indicates that HydA emerged within bacteria most likely from a Nar1-like ancestor lacking the 2Fe subcluster, and that this was followed by acquisition in several unicellular eukaryotes.
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Validity of average, minimum, and maximum end-of-day recall assessments of pain and fatigue.
Contemp Clin Trials
PUBLISHED: 02-24-2010
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End-of-day (EOD) diary assessments of symptoms have the potential to reduce recall bias associated with longer recall periods, and therefore be useful for generating accurate patient reported outcomes (PROs). In this report we examine the relative validity of diary questions about the experience of daily pain and fatigue, including several questions about experience for the entire day and questions about minimum and maximum daily levels, with previously collected data. Validity estimates are based on comparisons of EOD reports with momentary recordings of pain and fatigue from the same days. One hundred and six participants with rheumatologic diseases yielded 2852 days for analysis. Differences in levels as assessed by EOD and momentary reports were small (just a few points), although in many instances were significantly different. Correlational analyses indicated that "how much," "how intense," and "on average" EOD questions were more strongly associated with momentary reports (rs=0.85-0.90 for pain and 0.81-0.83 for fatigue) than were minimum and maximum questions (rs=0.73-0.80 for pain and 0.67-0.75 for fatigue). Overall, the pain measures had higher EOD-momentary correspondence than the fatigue measures. Analyses of difference scores between EOD and momentary reports confirmed the better correspondence of the average questions compared with minimum and maximum questions. There was little evidence of individual differences in level and correspondence analyses. The implication of these results is that over-the-day diary measures may yield superior PROs than those based on minimum or maximum daily levels.
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The antiviral protein viperin is a radical SAM enzyme.
FEBS Lett.
PUBLISHED: 02-08-2010
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Viperin, an interferon-inducible antiviral protein, is shown to bind an iron-sulfur cluster, based on iron analysis as well as UV-Vis and electron paramagnetic resonance spectroscopic data. The reduced protein contains a [4Fe-4S](1+) cluster whose g-values are altered upon addition of S-adenosylmethionine (SAM), consistent with SAM coordination to the cluster. Incubation of reduced viperin with SAM results in reductive cleavage of SAM to produce 5-deoxyadenosine (5-dAdo), a reaction characteristic of the radical SAM superfamily. The 5-dAdo cleavage product was identified by a combination of HPLC and mass spectrometry analysis.
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Complete stereospecific repair of a synthetic dinucleotide spore photoproduct by spore photoproduct lyase.
J. Biol. Inorg. Chem.
PUBLISHED: 01-04-2010
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Spore photoproduct lyase (SP lyase), a member of the radical S-adenosylmethionine superfamily of enzymes, catalyzes the repair of 5-thyminyl-5,6-dihydrothymine [spore photoproduct (SP)], a type of UV-induced DNA damage unique to bacterial spores. The anaerobic purification and characterization of Clostridium acetobutylicum SP lyase heterologously expressed in Escherichia coli, and its catalytic activity in repairing stereochemically defined synthetic dinucleotide SPs was investigated. The purified enzyme contains between 2.3 and 3.1 iron atoms per protein. Electron paramagnetic resonance (EPR) spectroscopy reveals an isotropic signal centered at g = 1.99, characteristic of a [3Fe-4S](+) cluster accounting for 3-4% of the iron in the sample. Upon reduction, a nearly axial signal (g = 2.03, 1.93 and 1.92) characteristic of a [4Fe-4S](+) cluster is observed that accounts for 34-45% of total iron. Addition of S-adenosylmethionine to the reduced enzyme produces a rhombic signal (g = 2.02, 1.93, 1.82) unique to the S-adenosyl-L: -methionine complex while decreasing the overall EPR intensity. This reduced enzyme is shown to rapidly and completely repair the 5R diastereomer of a synthetic dinucleotide SP with a specific activity of 7.1 +/- 0.6 nmol min(-1) mg(-1), whereas no repair was observed for the 5S diastereomer.
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Identification and characterization of a novel member of the radical AdoMet enzyme superfamily and implications for the biosynthesis of the Hmd hydrogenase active site cofactor.
J. Bacteriol.
PUBLISHED: 11-06-2009
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The genetic context, phylogeny, and biochemistry of a gene flanking the H(2)-forming methylene-H(4)-methanopterin dehydrogenase gene (hmdA), here designated hmdB, indicate that it is a new member of the radical S-adenosylmethionine enzyme superfamily. In contrast to the characteristic CX(3)CX(2)C or CX(2)CX(4)C motif defining this family, HmdB contains a unique CX(5)CX(2)C motif.
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The iron-sulfur cluster of pyruvate formate-lyase activating enzyme in whole cells: cluster interconversion and a valence-localized [4Fe-4S]2+ state.
Biochemistry
PUBLISHED: 08-29-2009
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Pyruvate formate-lyase activating enzyme (PFL-AE) catalyzes the generation of a catalytically essential glycyl radical on pyruvate formate-lyase (PFL). Purified PFL-AE contains an oxygen-sensitive, labile [4Fe-4S] cluster that undergoes cluster interconversions in vitro, with only the [4Fe-4S](+) cluster state being catalytically active. Such cluster interconversions could play a role in regulating the activity of PFL-AE, and thus of PFL, in response to oxygen levels in vivo. Here we report a Mossbauer investigation on whole cells overexpressing PFL-AE following incubation under aerobic and/or anaerobic conditions and provide evidence that PFL-AE undergoes cluster interconversions in vivo. After 2 h aerobic induction of PFL-AE expression, approximately 44% of the total iron is present in [4Fe-4S](2+) clusters, 6% in [2Fe-2S](2+) clusters, and the remainder as noncluster Fe(III) (29%) and Fe(II) (21%) species. Subsequent anaerobic incubation of the culture results in approximately 75% of the total iron being present as [4Fe-4S](2+) clusters, with no detectable [2Fe-2S](2+). Ensuing aerobic incubation of the culture converts the iron species nearly back to the original composition (42% [4Fe-4S](2+), 10% [2Fe-2S](2+), 19% Fe(III), and 29% Fe(II)). The results provide evidence for changes in cluster composition of PFL-AE in response to the redox state of the cell. Furthermore, the Mossbauer spectra reveal that the [4Fe-4S](2+) cluster of PFL-AE in whole cells contains a valence-localized Fe(III)Fe(II) pair which has not been previously observed in the purified enzyme. Addition of certain small molecules containing adenosyl moieties, including 5-deoxyadenosine, AMP, ADP, and methylthioadenosine, to purified PFL-AE reproduces the valence-localized state of the [4Fe-4S](2+) cluster. It is speculated that the [4Fe-4S](2+) cluster of PFL-AE in whole cells may be coordinated by a small molecule, probably AMP, and that such coordination may protect this labile cluster from oxidative damage.
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Chemoselective deprotection of triethylsilyl ethers.
Nucleosides Nucleotides Nucleic Acids
PUBLISHED: 08-28-2009
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An efficient and selective method was developed for the deprotection of triethylsilyl (TES) ethers using formic acid in methanol (5-10%) or in methylene chloride 2-5%) with excellent yields. TES ethers are selectively deprotected to the corresponding alcohols in high yields using formic acid in methanol under mild reaction conditions. Other hydroxyl protecting groups like t-butyldimethylsilyl (TBDMS) remain unaffected.
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Activation of HydA(DeltaEFG) requires a preformed [4Fe-4S] cluster.
Biochemistry
PUBLISHED: 05-14-2009
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The H-cluster is a complex bridged metal assembly at the active site of [FeFe]-hydrogenases that consists of a [4Fe-4S] subcluster bridged to a 2Fe-containing subcluster with unique nonprotein ligands, including carbon monoxide, cyanide, and a dithiolate ligand of unknown composition. Specific biosynthetic gene products (HydE, HydF, and HydG) responsible for the biosynthesis of the H-cluster and the maturation of active [FeFe]-hydrogenase have previously been identified and shown to be required for the heterologous expression of active [FeFe]-hydrogenase [Posewitz, M. C., et al. (2004) J. Biol. Chem. 279, 25711-25720]. The precise roles of the maturation proteins are unknown; the most likely possibility is that they are directed at the synthesis of the entire 6Fe-containing H-cluster, the 2Fe subcluster, or only the unique ligands of the 2Fe subcluster. The spectroscopic and biochemical characterization of HydA(DeltaEFG) (the [FeFe]-hydrogenase structural protein expressed in the absence of the maturation machinery) reported here indicates that a [4Fe-4S] cluster is incorporated into the H-cluster site. The purified protein in a representative preparation contains Fe (3.1 +/- 0.5 Fe atoms per HydA(DeltaEFG)) and S(2-) (1.8 +/- 0.5 S(2-) atoms per HydA(DeltaEFG)) and exhibits UV-visible spectroscopic features characteristic of iron-sulfur clusters, including a bleaching of the visible chromophore upon addition of dithionite. The reduced protein gave rise to an axial S = (1)/(2) EPR signal (g = 2.04 and 1.91) characteristic of a reduced [4Fe-4S](+) cluster. Mossbauer spectroscopic characterization of (57)Fe-enriched HydA(DeltaEFG) provided further evidence of the presence of a redox active [4Fe-4S](2+/+) cluster. Iron K-edge EXAFS data provided yet further support for the presence of a [4Fe-4S] cluster in HydA(DeltaEFG). These spectroscopic studies were combined with in vitro activation studies that demonstrate that HydA(DeltaEFG) can be activated by the specific maturases only when a [4Fe-4S] cluster is present in the protein. In sum, this work supports a model in which the role of the maturation machinery is to synthesize and insert the 2Fe subcluster and/or its ligands and not the entire 6Fe-containing H-cluster bridged assembly.
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Can End-of-day reports replace momentary assessment of pain and fatigue?
J Pain
PUBLISHED: 05-14-2009
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This study evaluated the ability of end-of-day (EOD) ratings to accurately reflect momentary (EMA) ratings on 10 widely used pain and fatigue items. Rheumatology patients (n = 105) completed >or=5 randomly scheduled EMA assessments of each item per day as well as EOD ratings. Correlations were high between EOD and EMA ratings of the 5 pain items (r = .90 to .92) and somewhat lower for the 5 fatigue/energy items (r = .71 to .86). To examine the ability of EOD ratings to represent 1 week of EMA ratings, 7 EOD ratings were averaged and correlated with EMA (r >or= .95 for pain items, r = .88 to .95 for fatigue/energy items). Further, averaging only 3 to 5 EOD ratings achieved very high correlations with 1 week of EMA ratings. Within-subject correlations of EOD with mean daily EMA across 7 days confirmed patients ability to provide daily ratings that accurately reflect their day-to-day variation in symptom levels. These EOD results were compared to traditional recall ratings collected in the same protocol. It was concluded (1) that EOD ratings were a better representation of EMA than were recall ratings, and (2) that EOD ratings across a reporting period can replace EMA for studies targeting average levels of pain or fatigue.
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Hydrogenase cluster biosynthesis: organometallic chemistry natures way.
Dalton Trans
PUBLISHED: 03-30-2009
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It has been over a decade now since it was revealed that the metal containing active sites of hydrogenases possess carbonyl and cyanide ligands bound to iron. The presence of these ligands in hydrogenases came as a surprise and to-date these ligands have not been observed to be associated with any other enzymatic metallocenter. The elucidation of the structures of these unique metalloenzymes and their associated metal clusters created opportunity for a number of different lines of research. For synthetic chemists, the structures of hydrogenase active sites have provided attractive targets for syntheses that advance our understanding of the electronic structure and reactivity of these unique enzyme active sites. These efforts contribute to the synthesis of first row transition metal catalysts for hydrogen oxidation and hydrogen production that could have significant impacts on alternative and renewable energy solutions. Although effective synthetic approaches have been identified to generate models with a high degree of similarity to these active sites, the details of how these metal clusters are synthesized biochemically have not been resolved. Since hydrogen metabolism is presumed to be an early feature in the energetics of life and hydrogen metabolizing organisms can be traced very early in molecular phylogeny, the metal clusters at hydrogenase active sites are presumed to be among the earliest of known co-factors. Comparison of mineral based precursors and synthetic cluster analog chemistry to what is observed in contemporary biological systems may shed light on how proto-metabolically relevant catalysts first arose prebiotically by the processes of adoption of pre-existing functionality and ligand assisted catalysis.
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Inspiratory airflow dynamics during sleep in irritable bowel syndrome: a pilot study.
Sleep Breath
PUBLISHED: 03-04-2009
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This study is a preliminary test of the hypothesis that the pathophysiology of irritable bowel syndrome (IBS) derives from pharyngeal collapse during sleep.
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Multi-modal examination of psychological and interpersonal distinctions among MPI coping clusters: a preliminary study.
J Pain
PUBLISHED: 02-26-2009
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The Multidimensional Pain Inventory (MPI) is a widely used instrument to characterize distinct psychosocial subgroups of patients with chronic pain: Adaptive (AC), Dysfunctional (DYS), and Interpersonally Distressed (ID). To date, several questions remain about the validity and distinctiveness of the patient clusters, and continued scientific attention has strongly been recommended. It is unclear if AC patients experience better adjustment or merely present themselves favorably. Moreover, differences in psychological distress and interpersonal relations between DYS and ID patients are equivocal. The present study is the first to utilize comprehensive informant ratings to extend prior validity research on the MPI. We employed a multimodal methodology consisting of patient self-report, parallel informant ratings, and behavioral measures. Ninety-nine patients with chronic pain, their partners, and providers participated. They completed measures of patients psychological distress and social relations. We also systematically observed patients pain behavior. Results provided strong support for the validity of the AC cluster in that patients positive adaptation was reliably corroborated by informants. The differentiating characteristics between the 2 maladaptive clusters, however, remain elusive. We found evidence that DYS patients distress appeared to be illness specific rather than generalized; however, both clusters were equally associated with social distress and partner/caregiver burden.
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Spore photoproduct lyase catalyzes specific repair of the 5R but not the 5S spore photoproduct.
J. Am. Chem. Soc.
PUBLISHED: 01-31-2009
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Bacterial spores are remarkable in their resistance to chemical and physical stresses, including exposure to UV radiation. The unusual UV resistance of bacterial spores is a result of the unique photochemistry of spore DNA, which results in accumulation of 5-thyminyl-5,6-dihydrothymine (spore photoproduct, or SP), coupled with the efficient repair of accumulated damage by the enzyme spore photoproduct lyase (SPL). SPL is a member of the radical AdoMet superfamily of enzymes, and utilizes an iron-sulfur cluster and S-adenosylmethionine to repair SP by a direct reversal mechanism initiated by H atom abstraction from C-6 of the thymine dimer. While two distinct diastereomers of SP (5R or 5S) could in principle be formed upon UV irradiation of bacterial spores, only the 5R configuration is possible for SP formed from adjacent thymines in double helical DNA, due to the constraints imposed by the DNA structure; the 5S configuration is possible in less well-defined DNA structures or as an interstrand cross-link. We report here results from HPLC and MS analysis of in vitro enzymatic assays on stereochemically defined SP substrates demonstrating that SPL specifically repairs only the 5R isomer of SP. The observation that 5R-SP, but not 5S-SP, is a substrate for SPL is consistent with the expectation that 5R is the SP isomer produced in vivo upon UV irradiation of bacterial spore DNA.
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Control of radical chemistry in the AdoMet radical enzymes.
Curr Opin Chem Biol
PUBLISHED: 01-30-2009
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The radical AdoMet superfamily comprises a diverse set of >2800 enzymes that utilize iron-sulfur clusters and S-adenosylmethionine (SAM or AdoMet) to initiate a diverse set of radical-mediated reactions. The intricate control these enzymes exercise over the radical transformations they catalyze is an amazing feat of elegance and sophistication in biochemistry. This review focuses on the accumulating evidence for how these enzymes control this remarkable chemistry, including controlling the reactivity between the iron-sulfur cluster and AdoMet, and controlling the subsequent radical transformations.
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Validation of the MPI patient profiles by partners and healthcare providers.
Pain
PUBLISHED: 01-11-2009
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The Multidimensional Pain Inventory (MPI) is one of the most widely used instruments to assess patients coping with chronic pain. It provides a psychosocial classification system that categorizes patients into three coping styles: Adaptive, Dysfunctional, and Interpersonally Distressed. To date, comprehensive information about the validity of the MPI taxonomy obtained from informants other than the patient has been unavailable. This has limited conclusions about the extent to which the MPI captures patients adaptation to chronic pain beyond self-report. The present study is the first to examine whether the distinct multidimensional profiles underlying the patient clusters can be confirmed by proxy report. Ninety-nine chronic pain patients, their partners, and their healthcare providers participated in the study. Patients completed the MPI twice to determine stability of classification. Partners and providers rated the patient on MPI proxy versions developed for this study. Results revealed that partner- and provider-reported MPI ratings corresponded with the self-report patient profiles. The profiles of patients showing classification stability rather than switching of cluster assignment between the two MPI assessments had the highest correspondence with proxy ratings. These results extend prior validity research on the MPI and demonstrate that differential psychological adaptational styles to chronic pain can be reliably recognized by partners and healthcare providers.
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Pittsburgh and Epworth sleep scale items: accuracy of ratings across different reporting periods.
Behav Sleep Med
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This study examined the ecological validity of sleep experience reports across different lengths of reporting periods. The accuracy of item responses on the Pittsburgh Sleep Quality Index (PSQI) and Epworth Sleepiness Scale (ESS) across 3-, 7-, and 28-day reporting periods was examined in relation to electronic daily item ratings. Primary care clinic patients (N = 119) were recruited, and were not required to have sleep problems to participate. Analyses found few differences in item scores when electronic daily ratings were compared with recall ratings, regardless of the length of the reporting period. However, within-subjects analyses indicated low levels of accuracy in recall of sleep items for specific days in the last week. Thus, for the purpose of between-subject comparisons, patients generally can provide accurate recall of sleep experiences; studies requiring finer-grained analysis across time and within-subjects require daily diary methodology.
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Emerging themes in radical SAM chemistry.
Curr. Opin. Struct. Biol.
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Enzymes in the radical SAM (RS) superfamily catalyze a wide variety of reactions through unique radical chemistry. The characteristic markers of the superfamily include a [4Fe-4S] cluster coordinated to the protein via a cysteine triad motif, typically CX(3)CX(2)C, with the fourth iron coordinated by S-adenosylmethionine (SAM). The SAM serves as a precursor for a 5-deoxyadenosyl radical, the central intermediate in nearly all RS enzymes studied to date. The SAM-bound [4Fe-4S] cluster is located within a partial or full triosephosphate isomerase (TIM) barrel where the radical chemistry occurs protected from the surroundings. In addition to the TIM barrel and a RS [4Fe-4S] cluster, many members of the superfamily contain additional domains and/or additional Fe-S clusters. Recently characterized superfamily members are providing new examples of the remarkable range of reactions that can be catalyzed, as well as new structural and mechanistic insights into these fascinating reactions.
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A challenging empirical question: What are the effects of media on psychogenic illness during a community crisis?
J Community Med Health Educ
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Psychogenic illness during disasters can cripple emergency healthcare services. Almost all research into this phenomenon has been retrospective and observational, and much of it suggests that media coverage can amplify psychogenic outbreaks. But there is little empirical evidence that this is true or that, conversely, media reports can mitigate psychogenic symptoms. In their work experimentally inducing psychogenic illness, the authors became sharply aware that it is difficult to experimentally mimic real-time media coverage. Yet clarifying medias effects on psychogenic illness is important if we want to prevent psychological disturbance. To meet this challenge, the authors advocate the funding and development of research protocols in advance of public emergencies, ready to be implemented in real-time. Coupled with digital media, which can track the reading and viewing behavior of millions of people, this approach can help us better understand medias impact on public health during an emergency, for better or for worse.
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Iron-sulfur cluster coordination in the [FeFe]-hydrogenase H cluster biosynthetic factor HydF.
FEBS Lett.
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Iron-sulfur cluster coordination was probed in the [FeFe]-hydrogenase H cluster maturation scaffold HydF. Putative Cys thiol and His imidazole ligation identified through multiple sequence alignments and structural studies were subjected to amino acid substitution and the variants were biochemically characterized. The results implicate a role for C304, C353, C356, and H306 of Clostridium acetobutylicum HydF in FeS cluster binding. Individual ligand substitutions affect both [4Fe-4S] and [2Fe-2S] cluster coordination suggesting shared coordination or cluster interconversion. Substitutions at C353 and H306 appear to preferentially impact the presence of the [2Fe-2S] cluster complement of the resulting variants of HydF. The results implicate a potential role for these residues in biosynthesis specifically and potential in bridging the [4Fe-4S] cluster to 2Fe subcluster biosynthetic intermediates.
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Radical SAM enzymes in methylation and methylthiolation.
Metallomics
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Radical S-adenosyl-l-methionine (SAM) enzymes are a large and diverse superfamily with functions ranging from enzyme activation through a single H atom abstraction to complex organic and metal cofactor synthesis involving a series of steps. Though these enzymes carry out a variety of functions, they share common structural and mechanistic characteristics. All of them contain a site-differentiated [4Fe-4S] cluster, ligated by a CX(3)CX(2)C or similar motif, which binds SAM at the unique iron. The [4Fe-4S](1+) state of the cluster reductively cleaves SAM to produce a 5-deoxyadenosyl radical, which serves to initiate the diverse reactions catalyzed by these enzymes. Recent highlights in the understanding of radical SAM enzymes will be presented, with a particular emphasis on enzymes catalyzing methylation and methythiolation reactions.
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Meeting them where they are: Using the Internet to deliver behavioral medicine interventions for pain.
Transl Behav Med
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Pharmacological and interventional pain medicine treatments are emphasized in the routine treatment of chronic pain despite strong evidence for the efficacy and safety of behavioral approaches. Most medical professionals have not incorporated behavioral pain treatments into their practices. Internet-based interventions have the potential to increase clinical use of these treatments. We discuss the strengths and weaknesses of current Internet-based behavioral pain management interventions, focusing on three broad intervention categories: therapist-guided interventions, unguided (automated) interventions, and pain-relevant applications for mobile platforms. Examples of each category are discussed, revealing a high degree of variation in approaches, user interfaces, and components as well as variability in the degree to which these interventions have been subjected to empirical testing. Finally, we highlight key issues for research and clinical implementation, with the goal of advancing this field so that it can meet its potential to increase access to evidence-based behavioral medicine treatments for chronic pain.
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Expanding options for developing outcome measures from momentary assessment data.
Psychosom Med
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We propose several different patient-reported outcomes (PROs) from momentary, real-time collection of symptom data. In addition to the mean of momentary reports of symptoms, other types of summaries can reflect different aspects of the symptom experience.
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Emerging paradigms for complex iron-sulfur cofactor assembly and insertion.
Annu. Rev. Biochem.
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[FeFe]-hydrogenses and molybdenum (Mo)-nitrogenase are evolutionarily unrelated enzymes with unique complex iron-sulfur cofactors at their active sites. The H cluster of [FeFe]-hydrogenases and the FeMo cofactor of Mo-nitrogenase require specific maturation machinery for their proper synthesis and insertion into the structural enzymes. Recent insights reveal striking similarities in the biosynthetic pathways of these complex cofactors. For both systems, simple iron-sulfur cluster precursors are modified on assembly scaffolds by the activity of radical S-adenosylmethionine (SAM) enzymes. Radical SAM enzymes are responsible for the synthesis and insertion of the unique nonprotein ligands presumed to be key structural determinants for their respective catalytic activities. Maturation culminates in the transfer of the intact cluster assemblies to a cofactor-less structural protein recipient. Required roles for nucleotide binding and hydrolysis have been implicated in both systems, but the specific role for these requirements remain unclear. In this review, we highlight the progress on [FeFe]-hydrogenase H cluster and nitrogenase FeMo-cofactor assembly in the context of these emerging paradigms.
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Individual differences in the day-to-day variability of pain, fatigue, and well-being in patients with rheumatic disease: associations with psychological variables.
Pain
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This report examines day-to-day variability in rheumatology patients ratings of pain and related quality-of-life variables as well as predictors of that variability. Data from 2 studies were used. The hypothesis was that greater psychological distress (i.e., depression and anxiety) and poorer coping appraisals (i.e., higher pain catastrophizing and lower self-efficacy) are associated with more variability. Electronic daily diary ratings were collected from 106 patients from a community rheumatology practice across 28 days (study 1) and from 194 osteoarthritis patients across 7 days (study 2). In multilevel modeling analyses, substantial day-to-day variability was evident for all variables in both studies, and individual patients differed considerably and somewhat reliably in the magnitude of their variability. Higher levels of depression significantly predicted greater variability in pain, as well as in happiness and frustration (study 1). Lower self-efficacy was associated with more variability in patients daily satisfaction with accomplishments and in the quality of their day (study 2). Greater pain catastrophizing and higher depression predicted more variability in interference with social relationships (study 2). Anxiety was not significantly associated with day-to-day variability. The results of these studies suggest that individual differences in the magnitude of symptom fluctuation may play a vital role in understanding patients adjustment to pain. Future research will be needed to examine the clinical utility of measuring variability in patients pain and well-being, and to understand whether reducing variability may be an important treatment target.
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Genome sequence of Desulfitobacterium hafniense DCB-2, a Gram-positive anaerobe capable of dehalogenation and metal reduction.
BMC Microbiol.
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The genome of the Gram-positive, metal-reducing, dehalorespiring Desulfitobacterium hafniense DCB-2 was sequenced in order to gain insights into its metabolic capacities, adaptive physiology, and regulatory machineries, and to compare with that of Desulfitobacterium hafniense Y51, the phylogenetically closest strain among the species with a sequenced genome.
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Radical AdoMet enzymes in complex metal cluster biosynthesis.
Biochim. Biophys. Acta
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Radical S-adenosylmethionine (AdoMet) enzymes comprise a large superfamily of proteins that engage in a diverse series of biochemical transformations through generation of the highly reactive 5-deoxyadenosyl radical intermediate. Recent advances into the biosynthesis of unique iron-sulfur (FeS)-containing cofactors such as the H-cluster in [FeFe]-hydrogenase, the FeMo-co in nitrogenase, as well as the iron-guanylylpyridinol (FeGP) cofactor in [Fe]-hydrogenase have implicated new roles for radical AdoMet enzymes in the biosynthesis of complex inorganic cofactors. Radical AdoMet enzymes in conjunction with scaffold proteins engage in modifying ubiquitous FeS precursors into unique clusters, through novel amino acid decomposition and sulfur insertion reactions. The ability of radical AdoMet enzymes to modify common metal centers to unusual metal cofactors may provide important clues into the stepwise evolution of these and other complex bioinorganic catalysts. This article is part of a Special Issue entitled: Radical SAM enzymes and Radical Enzymology.
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Obesity and pain are associated in the United States.
Obesity (Silver Spring)
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Recent small-scale studies have shown a positive association between central obesity and self-reported pain levels. This study attempts to replicate the finding in a survey of over 1,000,000 individuals in the United States. The Gallup Organization conducted a proprietary survey between 2008 through 2010 where 1,062,271 randomly selected individuals in the United States participated in a telephone interview. Survey questions included height and weight, from which BMI was computed, questions about pain conditions in the past year, and a question about pain experience yesterday. Only 36.8% [corrected] of the sample was classified as Low-Normal BMI, 38.6 [corrected] were classified as Overweight, and the remainder was in the three categories of Obese. BMI and pain yesterday were reliably associated when demographic variables were controlled: the overweight group reported 20% higher rates of pain than Low-Normal group, 68% higher for Obese I group, 136% higher for Obese II group, and 254% higher for Obese III group. The association held for both men and women and it became stronger in older age groups. Controlling the associations for other pain-related medical conditions substantially reduced the associations, but they remained substantial for the Obese groups. We conclude that BMI and daily pain are positively correlated in the United States: people who are obese are considerably more prone to having daily pain. The association is robust and holds after controlling for several pain conditions and across gender and age. The increasing BMI-pain association with older ages suggests a developmental process that, along with metabolic hypotheses, calls out for investigation.
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Single-day Pain Assessments as Clinical Outcomes: Not So Fast.
Clin J Pain
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Pain diaries are important tools for clinical trials and optimal assay sensitivity of outcomes derived from diaries is a worthwhile goal. Jensen and colleagues recently reported results suggesting that single-day diary-based outcomes could possibly be as psychometrically sound as outcomes based on taking the average of many diaries.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.