In JoVE (1)
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Articles by Danu S. Perumalla 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.
Other articles by Danu S. Perumalla on PubMed
HyCCAPP As a Tool to Characterize Promoter DNA-protein Interactions in Saccharomyces Cerevisiae Genomics. 06, 2016 | Pubmed ID: 27184763 Currently available methods for interrogating DNA-protein interactions at individual genomic loci have significant limitations, and make it difficult to work with unmodified cells or examine single-copy regions without specific antibodies. In this study, we describe a physiological application of the Hybridization Capture of Chromatin-Associated Proteins for Proteomics (HyCCAPP) methodology we have developed. Both novel and known locus-specific DNA-protein interactions were identified at the ENO2 and GAL1 promoter regions of Saccharomyces cerevisiae, and revealed subgroups of proteins present in significantly different levels at the loci in cells grown on glucose versus galactose as the carbon source. Results were validated using chromatin immunoprecipitation. Overall, our analysis demonstrates that HyCCAPP is an effective and flexible technology that does not require specific antibodies nor prior knowledge of locally occurring DNA-protein interactions and can now be used to identify changes in protein interactions at target regions in the genome in response to physiological challenges.