X-ray crystal structures of rabbit N-acetylglucosaminyltransferase I (GnT I) in complex with donor substrate analogues.
The Golgi-resident glycosyltransferase, UDP-N-acetyl-d-glucosamine:alpha-3-d-mannoside beta-1,2-N-acetylglucosaminyltransferase I (GnT I), initiates the conversion of high-mannose oligosaccharides to complex and hybrid structures in the biosynthesis of N-linked glycans. Reported here are the X-ray crystal structures of GnT I in complex with UDP-CH2-GlcNAc (a non-hydrolyzable C-glycosidic phosphonate), UDP-2-deoxy-2-fluoro-glucose, UDP-glucose and UDP. Collectively, these structures provide evidence for the importance of the GlcNAc moiety and its N-acetyl group in donor substrate binding, as well as insight into the role played by the flexible 318-330 loop in substrate binding and product release. In addition, the UDP-CH2-GlcNAc complex reveals a well-defined glycerol molecule poised for nucleophilic attack on the C1 atom of the donor substrate analogue. The position and orientation of this glycerol molecule have allowed us to model the binding of the Manalpha1,3Manbeta1 moiety of the acceptor substrate and, based on the model, to suggest a rationalization for the main determinants of GnT I acceptor specificity.
Gordon
Roni D
RD
Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada M5S 1A8.
Sivarajah
Prashanth
P
Satkunarajah
Malathy
M
Ma
Dengbo
D
Tarling
Chris A
CA
Vizitiu
Dragos
D
Withers
Stephen G
SG
Rini
James M
JM
D000818:Animals; D002384:Catalysis; D018360:Crystallography, X-Ray; D016695:Glycosyltransferases / Q000737:chemistry; D007700:Kinetics; D008358:Mannose / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011817:Rabbits; D013379:Substrate Specificity
D009844:Oligosaccharides; D016695:Glycosyltransferases; D017351:N-Acetylglucosaminyltransferases; C024850:alpha-1,3-mannosyl-glycoprotein beta-1,2-N-acetylglucosaminyltransferase I; D008358:Mannose