POSTTRANSLATIONAL MODIFICATIONS OF CYTOKERATIN PROTEINS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our goal is to determine phosphorylation and O-linked glycosylation sites on cytokeratin polypeptide 8, studying cytokeratin polypeptide 8 and possibly cytokeratin polypeptide 18 at the mass spectrometry level. We hope to obtain detailed structural information to support our initial biochemical data indicating that this cytokeratin has a single N-acetylglucosamine residue we presume is linked to serine. We also hope our system coupled with mass spectrometry would determine if there is an inverse relationship between the state of serine phosphorylation and O-linked glycosylation. Cytokeratin proteins make up an ideal system to study this question since they are heavily phosphorylated on serine residues and in addition have the O-linked glycosylation and are present in cells in large quantities, making them amenable for mass spec sequencing studies. A variety of methods are being pursued aimed at development of a robust mass spectrometric strategy for the site specific characterization of O-GlcNAc residues on nuclear proteins for comparison with phosphorylation sites on the same proteins.. 293C