SEPARATION OF HISTONES & CHARC OF INTACT PROTEINS &MODIFICATIONS BY MASS SPEC

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Histones are a group of highly conserved proteins wrapped tightly by DNA in chromatin nucleosomes. They are rich in modifications such as acetylation, methylation, phosphorylation, mono-ubiquitylation, glycosylation, and ADP that influence the accessibility of DNA, its architecture and functions. Knowledge about these post-translational modifications in histones is vital to understanding the molecular processes involving DNA. The variety of modifications present in the various families of histones makes global analysis difficult and time consuming. Therefore, simple and reliable methods capable of their separating the various isoforms are desirable to aid in the elucidation of their modifications. This research project is directed at the development of methods for the separation of the various types of histones and the isoforms within each type to reduce sample complexity. This will be achieved using a combination of gel electrophoresis, reverse phase and HILIC chromatography. It is hoped that these simplified samples will enable the identification of post-translational modifications within each histone by FT-ICR or Orbitrap MS.