Identification of Secondary Structure in Peptides From Mucin Protein Involved in UDP-GALNAC:Polypeptide Nacetylgalactosaminyltransferase
Mucins, a protein component of mucus, have various cell surface functions. They provide resistance, pathogen interference, and mediate cell-cell interaction. Cells over express mucin as a response to avoid destruction. The viscosity, due to the abundance of saccharide residues, provides an impenetrable coating. This particular immune system response is regulated by glycosylation. The bulk of mucin is formed from galactose. Glycosylation catalyzed by the enzyme UDP-GalNAc:Polypeptide NAcetylgalactosaminyltransferase (GalNAcT) elongates mucin by adding galactose. The 800 residue protein only has 196 sites that recognize (GalNAcT) but only four sites can transfer galactose. My project is a continuation from previous studies of Ake Elhammer (1993). Serine and threonine on the protein surface were found to have the best enzyme binding rate. Another study was done by Elhammer (1995) on acceptor sequences that do no contain a serine or threonine. These sequences would bind the enzyme but no galactose was transferred. Since a specific amino acid residue was not solely responsible for acceptable binding strengths, structure in relation to accessibility of these particular protein surface acceptor sites was investigated. We analyzed the strongest binding peptide, U-109367, and three with similar structure, U-96663, U-140614, and U-140615, that bind to UDPGalNAc: Polypeptide N-Acetylgalactoseaminyltransferase. Data was taken at several temperatures of each of the peptides to examine structure stability. The proton spin systems were examined by 2-D 1H COSY and ROESY nuclear magnetic resonance spectroscopy. Any interactions between hydrogens of adjacent bonds on an amino acid and of side chains on neighboring peptide residues within 5.0 A allows us to determine the secondary structure. Structural differences were found for each of the four peptides and at the various temperatures.