Structure-Based Mechanism and Specificity of Human Galactosyltransferase β3GalT5

• Published in: Journal of the American Chemical Society Vol. 147; no. 13; pp. 10875 - 10885
• Main Authors: Lo, Jennifer M., Kung, Chih-Chuan, Cheng, Ting-Jen Rachel, Wong, Chi-Huey, Ma, Che
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 02.04.2025
• Subjects: Crystallography, X-Ray; drug design; enzymes; galactose; Galactosyltransferases - chemistry; Galactosyltransferases - metabolism; glycolipids; glycoproteins; glycosylation; Humans; Models, Molecular; neoplasm progression; neoplasms; polysaccharides; Substrate Specificity; X-radiation
• ISSN: 0002-7863; 1520-5126; 1520-5126
• DOI: 10.1021/jacs.4c11724

Human β1,3-galactosyltransferase 5 (β3GalT5) is a key enzyme involved in the synthesis of glycans on glycoproteins and glycolipids that are associated with various important biological functions, especially tumor malignancy and cancer progression, and has been considered as a promising target for development of anticancer agents. In this study, we determined the X-ray structures of β3GalT5 in complex with the stable donor analogue UDP-2-fluorogalactose or the native donor substrate UDP-galactose (UDP-Gal) and several glycan acceptors at different reaction steps. Based on the structures obtained from our experiments, β3GalT5 catalyzes the transfer of galactose from UDP-Gal to a broad spectrum of glycan acceptors with an SN2-like mechanism; however, in the absence of a glycan acceptor, UDP-Gal is slowly converted to UDP and two other products, one is galactose through an SN2-like mechanism with water as an acceptor and the other is an oxocarbenium-like product, presumably through an SN1-like mechanisms. The structure, mechanism, and specificity of β3GalT5 presented in this study advance our understanding of enzymatic glycosylation and provide valuable insights for application to glycan synthesis and drug design targeting β3GalT5-associated cancer.