Sequences in the NCAM Ig5-‐FN1 Tandem Affect Interactions with the Polysialyltransferases
thesisposted on 22.02.2015 by Matthew G. Thompson
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Sequences in the NCAM Ig5-FN1 Tandem Affect Interactions with the Polysialyltransferases Matthew Gregory Thompson, Ph.D. Department of Biochemistry and Molecular Genetics University of Illinois at Chicago Chicago, Illinois (2012) Dissertation Chairperson: Karen J. Colley, Ph. D. The Neural Cell Adhesion Molecule (NCAM) carries a unique post-translational modification, polysialic acid, which modulates the its adhesive and signaling functions and consequently plays roles in nervous system development and plasticity, neuronal regeneration and cancer metastasis. The addition of polysialic acid is carried out by the polysialyltransferases ST8SiaII/STX and ST8SiaIV/PST. Polysialic acid is found on a limited number of proteins and previous work indicates that its addition requires a protein-protein interaction between the polysialyltransferase and substrate prior to modification of the substrate glycans. To evaluate the basis for this unique specificity, the interaction between NCAM and the polysialyltransferases was examined in this work. In this study I investigated sequences in NCAM’s first fibronectin type III repeat (FN1) and fifth immunoglobulin like (Ig5) domain and their role in polyST recognition and interaction. Results from these investigations revealed key residues in both the FN1 and Ig5 domain that affects the ability of the polysialyltransferases to interact with NCAM to allow subsequent NCAM polysialylation. Sequences in FN1, most notably a surface acidic patch, are involved in the interaction of the polyST with NCAM. Sequences at the interface of the FN1 and Ig5 domain are critical in maintaining the Ig5-FN1 relationship to allow NCAM polysialylation. Finally using a non-polysialylated NCAM homolog, the Olfactory Cell Adhesion Molecule (OCAM) to create chimeric proteins, and evaluating their polysialylation, revealed that specific residues in OCAM Ig5 domain block its polysialylation and the analogous residues in NCAM Ig5 are permissive for polysialylation. Taken together these results demonstrate for the first time that the polysialyltransferases recognize NCAM by interacting with residues in both the Ig5 and FN1 domains, and suggest new strategies for controlling the extent of NCAM polysialylation during development and in disease.