Mucin type O-glycosylation is an essential post translational protein modification linked to multiple disease states including many cancers and is critical to development. Our lab studies the control of mucin type O-glycosylation at the transferase and peptide acceptor substrate level. We utilize enzyme activity/kinetics and structural biological approaches to deduce how the initiating glycosyltransferases select and recognize specific sites to glycosylate in protein substrates.
Research Information
Research Interests
Biosynthesis of O-Linked Glycans
Glycoproteins containing heavily O-glycosylated, mucin-like, domains play important and diverse biological roles for example, protecting cell surfaces, modulating cell-cell interactions, targeting cellular proteins, regulating inflammatory and immune responses and in tumorogenisis and metastasis. In these glycoproteins O-glycosylated domains typically play critical functional roles, which rely on their extended structures and ability to be decorated by an array of glycan structures. The goals of our laboratory are to develop a detailed understanding of biosynthetic processes that regulate the unique glycosylation patterns of these heavily O-glycosylated domains. From these studies we will learn how O-glycosylation is modulated by peptide sequence and neighboring glycosylation and the extent that changes in glycosylation may alter the properties of these domains.
A very large family of ppGalNAc transferases (>16) initiates mucin-type O-glycan by adding N-acetylgalactosamine (GalNAc) to peptide Ser and Thr residues. Homologous family members are found over a range of species suggesting that certain ppGalNAc transferases may have unique substrate specificities evolutionarily maintained for the glycosylation of specific peptide sequences. Presently, the peptide substrate specificities of the ppGalNAc transferase family members are poorly characterized, leading to an inability to rationally predict or comprehend O-glycosylation. Our laboratory has developed several novel methods utilizing mucin derived peptides and random peptide substrates for evaluating the substrate specificity of these transferases. We have further developed a kinetic modeling approach capable of approximating the observed glycosylation patterns of the mucin peptide substrates. We have also shown that our modeling approach can also reproduce the in vivo substitution pattern of the peptide linked GalNAc by galactose, forming the so-called Core 1 base structure. With a fundamental understanding of ppGalNAc transferase specificity the identification of isoform-specific substrates, the creation of isoform-specific inhibitors, and the prediction of O-glycosylation sites will be made possible.
Research Projects
Ongoing studies in the Gerken lab include the comparison of ppGalNAc T specificity across species, the determination of glycopeptide substrate preferences and a collaboration with Dr. Stanford Markowitz at CWRU characterizing the properties and biological roles of a series of ppGalNAc Ts that are associated with human colon cancers. Studies are also in progress characterizing the substrate specificity of the purified Core 1 transferase along with several other GalNAc substituting transferases.
As a result of our work, a more complete understanding of the initial steps in the biosynthesis of O-glycans will be obtained. It is anticipated that these studies will lead to development of methods for the prediction of O-glycan structures in a site-specific manner. This work is funded by the National Cancer Institute of the National Institutes of Health and the Cystic Fibrosis Foundation.
Publications
- Sletmoen M., Gerken T. A., Stokke B. T., Burchell J., and Brewer C. F.
“Tn and STn are members of a family of carbohydrate tumor antigens that possess carbohydrate-carbohydrate interactions”
Glycobiology 28 (7): 437-42 (2018). - Evans D. R., Venkitachalam S., Revoredo L., Dohey A. T., Clarke E., Pennell J. J., Powell A. E., Quinn E., Ravi L., Gerken T. A., Green J. S., Woods M. O., and Guda K.
“Evidence for GALNT12 as a moderate penetrance gene for colorectal cancer”
Hum Mutat 39 (8): 1092-101 (2018). - de Las Rivas M., Lira-Navarrete E., Daniel E. J. P., Companon I., Coelho H., Diniz A., Jimenez-Barbero J., Peregrina J. M., Clausen H., Corzana F., Marcelo F., Jimenez-Oses G., Gerken T. A., and Hurtado-Guerrero R.
“The interdomain flexible linker of the polypeptide GalNAc transferases dictates their long-range glycosylation preferences”
Nat Commun 8 (1): 1959 (2017). - Revoredo L., Wang S., Bennett E. P., Clausen H., Moremen K. W., Jarvis D. L., Ten Hagen K. G., Tabak L. A., and Gerken T. A.
“Mucin-type O-glycosylation is controlled by short- and long-range glycopeptide substrate recognition that varies among members of the polypeptide GalNAc transferase family”
Glycobiology 26 (4): 360-76 (2016). - Haugstad K. E., Hadjialirezaei S., Stokke B. T., Brewer C. F., Gerken T. A., Burchell J., Picco G., and Sletmoen M.
“Interactions of mucins with the Tn or Sialyl Tn cancer antigens including MUC1 are due to GalNAc-GalNAc interactions”
Glycobiology 26 (12): 1338-50 (2016). - Gerken T. A., Revoredo L., Thome J. J., Tabak L. A., Vester-Christensen M. B., Clausen H., Gahlay G. K., Jarvis D. L., Johnson R. W., Moniz H. A., and Moremen K.
“The lectin domain of the polypeptide GalNAc transferase family of glycosyltransferases (ppGalNAc Ts) acts as a switch directing glycopeptide substrate glycosylation in an N- or C-terminal direction, further controlling mucin type O-glycosylation”
J Biol Chem 288 (27): 19900-14 (2013). - Gerken T. A., Jamison O., Perrine C. L., Collette J. C., Moinova H., Ravi L., Markowitz S. D., Shen W., Patel H., and Tabak L. A.
“Emerging paradigms for the initiation of mucin-type protein O-glycosylation by the polypeptide GalNAc transferase family of glycosyltransferases”
J Biol Chem 286 (16): 14493-507 (2011).