This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Tetrahymena thermophila encodes a histone 2A variant protein (HV1) that is studied by Dr. David Allis and colleagues at The Rockefeller University. They have observed variable modifications on this protein with directed antibodies, which change under different growth conditions. Of particular interest, the Allis group was interested in the extent of acetylation in normal growth conditions verses starvation conditions. HV1 was purified from T. thermophila that was grown under the two former conditions. We performed MALDI-TOF analysis of the intact protein to probe for post-translational modifications and specifically acetylation. We observed up to 5 acetyl groups and an average level of 2 acetyl groups on HV1 under normal growth conditions, while up to 3 acetyl groups and an average of 0-1 acetyl groups were detected under starvation conditions. The reduction of acetylation is consistent with condensed chromatin, which may be present in starved cells that are impaired in growth. Current efforts in this work include (1) determining the sites of acetylation by utilizing our MALDI-ion trap mass spectrometer to fragment tryptic peptides to probe for specific sites of acetylation (2) analyze all tryptic peptides for other post-translational modifications by mass analysis and peptide fragmentation.
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