isolation-lipoprotein-particles-from-chicken-egg-yolk-for-study

Isolation of Lipoprotein Particles from Chicken Egg Yolk for the Study of Bacterial Pathogen Fatty Acid Incorporation into Membrane Phospholipids

Staphylococcus aureus and other Gram-positive pathogens incorporate fatty acids from the environment into membrane phospholipids. During infection, the ...

Michigan State University

Staphylococcus aureus is the leading cause of skin and soft tissue infections, bacteremia, osteomyelitis, and endocarditis in the developed world. The ability of S. aureus to cause substantial disease in distinct host environments is supported by a flexible metabolism that allows this pathogen to overcome challenges unique to each host organ. One feature of staphylococcal metabolic flexibility is a branched aerobic respiratory chain composed of multiple terminal oxidases. Whereas previous biochemical and spectroscopic studies reported the presence of three different respiratory oxygen reductases (o type, bd type, and aa3 type), the genome contains genes encoding only two respiratory oxygen reductases, cydAB and qoxABCD Previous investigation showed that cydAB and qoxABCD are required to colonize specific host organs, the murine heart and liver, respectively. This work seeks to clarify the relationship between the genetic studies showing the unique roles of the cydAB and qoxABCD in virulence and the respiratory reductases reported in the literature. We establish that QoxABCD is an aa3-type menaquinol oxidase but that this enzyme is promiscuous in that it can assemble as a bo3-type menaquinol oxidase. However, the bo3 form of QoxABCD restricts the carbon sources that can support the growth of S. aureus In addition, QoxABCD function is supported by a previously uncharacterized protein, which we have named CtaM, that is conserved in aerobically respiring Firmicutes In total, these studies establish the heme A biosynthesis pathway in S. aureus, determine that QoxABCD is a type aa3 menaquinol oxidase, and reveal CtaM as a new protein required for type aa3 menaquinol oxidase function in multiple bacterial genera.

CtaM Is Required for Menaquinol Oxidase aa3 Function in Staphylococcus aureus.

Methicillin-resistant  (MRSA) is a threat to global health. Consequently, much effort has focused on the development of new antimicrobials that target novel aspects of  physiology. Fatty acids are required to maintain cell viability, and bacteria synthesize fatty acids using the type II fatty acid synthesis (FASII) pathway. FASII is significantly different from human fatty acid synthesis, underscoring the therapeutic potential of inhibiting this pathway. However, many Gram-positive pathogens incorporate exogenous fatty acids, bypassing FASII inhibition and leaving the clinical potential of FASII inhibitors uncertain. Importantly, the source(s) of fatty acids available to pathogens within the host environment remains unclear. Fatty acids are transported throughout the body by lipoprotein particles in the form of triglycerides and esterified cholesterol. Thus, lipoproteins, such as low-density lipoprotein (LDL), represent a potentially rich source of exogenous fatty acids for  during infection. We sought to test the ability of LDLs to serve as a fatty acid source for  and show that cells cultured in the presence of human LDLs demonstrate increased tolerance to the FASII inhibitor triclosan. Using mass spectrometry, we observed that host-derived fatty acids present in the LDLs are incorporated into the staphylococcal membrane and that tolerance to triclosan is facilitated by the fatty acid kinase A, FakA, and Geh, a triacylglycerol lipase. Finally, we demonstrate that human LDLs support the growth of  fatty acid auxotrophs. Together, these results suggest that human lipoprotein particles are a viable source of exogenous fatty acids for  during infection. Inhibition of bacterial fatty acid synthesis is a promising approach to combating infections caused by  and other human pathogens. However,  incorporates exogenous fatty acids into its phospholipid bilayer. Therefore, the clinical utility of targeting bacterial fatty acid synthesis is debated. Moreover, the fatty acid reservoir(s) exploited by  is not well understood. Human low-density lipoprotein particles represent a particularly abundant  source of fatty acids and are present in tissues that  colonizes. Herein, we establish that  is capable of utilizing the fatty acids present in low-density lipoproteins to bypass both chemical and genetic inhibition of fatty acid synthesis. These findings imply that  targets LDLs as a source of fatty acids during pathogenesis.

Staphylococcus aureus Utilizes Host-Derived Lipoprotein Particles as Sources of Fatty Acids.

Pathogens have evolved elegant mechanisms to acquire essential nutrients from host environments. Sulfur is a requirement for bacterial growth and inorganic and organic sulfur-containing metabolites are abundant within the host-pathogen interface. A growing body of evidence suggests that pathogens are capable of scavenging both types of sulfur sources to fulfill the nutritional requirement. While therapeutic strategies focusing on inhibiting inorganic sulfate assimilation and cysteine synthesis show promise in vitro, in vivo efficacy maybe limited due to the diversity of host-derived sulfur sources and the fact that most pathogens are capable of acquiring multiple sources of sulfur.

Article	Year
CtaM Is Required for Menaquinol Oxidase aa3 Function in Staphylococcus aureus. mBio\| PubMed ID: 27406563	2016
Staphylococcus aureus Utilizes Host-Derived Lipoprotein Particles as Sources of Fatty Acids. Journal of bacteriology\| PubMed ID: 29581406	2018
Nutrient sulfur acquisition strategies employed by bacterial pathogens. Current opinion in microbiology\| PubMed ID: 30530037	2018

Neal D. Hammer

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