3 articles published in JoVE
Adaptation of Hybridization Capture of Chromatin-associated Proteins for Proteomics to Mammalian Cells Hector Guillen-Ahlers1,2, Prahlad K. Rao1, Danu S. Perumalla1, Maria J. Montoya1, Avinash Y.L. Jadhav1, Michael R. Shortreed3, Lloyd M. Smith3, Michael Olivier1,2 1Department of Genetics, Texas Biomedical Research Institute, 2Department of Internal Medicine-Molecular Medicine, Wake Forest University School of Medicine, 3Department of Chemistry, University of Wisconsin This is a method to identify novel DNA-interacting proteins at specific target loci, relying on sequence-specific capture of crosslinked chromatin for subsequent proteomic analyses. No prior knowledge about potential binding proteins, nor cell modifications are required. Initially developed for yeast, the technology has now been adapted for mammalian cells.
A G-quadruplex DNA-affinity Approach for Purification of Enzymatically Active G4 Resolvase1 Eric D. Routh*1, Steven D. Creacy*2, Peter E. Beerbower3, Steven A. Akman4, James P. Vaughn1, Philip J. Smaldino3 1Department of Cancer Biology, Wake Forest School of Medicine, 2YX Genomics, 3Department of Biology, Ball State University, 4Department of Hematology and Oncology, Roper St. Francis Hospital G4 Resolvase1 binds to G-quadruplex (G4) structures with the tightest reported affinity for a G4-binding protein and represents the majority of the G4-DNA unwinding activity in HeLa cells. We describe a novel protocol that harnesses the affinity and ATP-dependent unwinding activity of G4-Resolvase1 to specifically purify catalytically active recombinant G4R1.
Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy Manish S. Bharadwaj1, Daniel J. Tyrrell1, Mary F. Lyles1, Jamehl L. Demons1, George W. Rogers2, Anthony J. A. Molina1 1Department of Internal Medicine, Section on Gerontology and Geriatric Medicine, Wake Forest School of Medicine, 2Seahorse Biosciences Methods for biopsy of Vastus lateralis, preparation of purified mitochondria, and respirometric profiling are described. The use of small muscle volume makes this technique suitable for clinical research applications.