Substrate Engagement and Catalytic Mechanisms of N-Acetylglucosaminyltransferase V

alpha-Mannoside beta-1,6-N-acetylglucosaminyltransferase V (MGATS) is a mammalian glycosyltransferase involved in complex TS N-glycan formation, which strongly drives cancer when overexpressed. Despite intense interest, the catalytic mechanism of MGATS is not known in detail, precluding therapeutic exploitation. We solved structures of MGATS complexed to glycosyl donor and acceptor ligands, revealing an unforeseen role for donor-induced loop rearrangements in controlling acceptor substrate engagement. QM/ MM metadynamics simulations of MGATS catalysis highlight the key assisting role of G1u297 and reveal considerable conformational distortions imposed upon the glycosyl donor during transfer. Detailed mechanistic characterization of MGATS will aid inhibitor development to correct cancer-associated N-glycosylation.