Cysteine-to-serine mutants dramatically reorder the active site of human ABO(H) blood group B glycosyltransferase without affecting activity: structural insights into cooperative substrate binding.

Journal of Molecular Biology
B SchumanS V Evans

Abstract

A common feature in the structures of GT-A-fold-type glycosyltransferases is a mobile polypeptide loop that has been observed to participate in substrate recognition and enclose the active site upon substrate binding. This is the case for the human ABO(H) blood group B glycosyltransferase GTB, where amino acid residues 177-195 display significantly higher levels of disorder in the unliganded state than in the fully liganded state. Structural studies of mutant enzymes GTB/C80S/C196S and GTB/C80S/C196S/C209S at resolutions ranging from 1.93 to 1.40 A display the opposite trend, where the unliganded structures show nearly complete ordering of the mobile loop residues that is lost upon substrate binding. In the liganded states of the mutant structures, while the UDP moiety of the donor molecule is observed to bind in the expected location, the galactose moiety is observed to bind in a conformation significantly different from that observed for the wild-type chimeric structures. Although this would be expected to impede catalytic turnover, the kinetics of the transfer reaction are largely unaffected. These structures demonstrate that the enzymes bind the donor in a conformation more similar to the dominant solution rotamer and facil...Continue Reading

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Citations

Mar 2, 2011·Biochemistry·Semi Park, Stephen J Lippard
Feb 22, 2012·Acta Crystallographica. Section D, Biological Crystallography·Asha R JohalStephen V Evans
Feb 9, 2011·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·B SchumanS V Evans
May 20, 2011·Current Opinion in Biotechnology·Aram ChangJon S Thorson
Mar 4, 2017·Chembiochem : a European Journal of Chemical Biology·Lena Lisbeth GrimmThomas Peters
Sep 22, 2017·Cellular and Molecular Life Sciences : CMLS·Deborah HarrusTuomo Glumoff

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