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A parsimonious mechanism of sugar dehydration by human GDP-mannose-4,6-dehydratase

DOI: 10.1021/acscatal.9b00064 DOI Help

Authors: Martin Pfeiffer (Graz University of Technology) , Catrine Johansson (Structural Genomics Consortium Oxford, University of Oxford; Botnar Research Centre, University of Oxford) , Tobias Krojer (Structural Genomics Consortium, University of Oxford) , Kathryn L. Kavanagh (Structural Genomics Consortium, University of Oxford) , Udo C. T. Oppermann (Structural Genomics Consortium, University of Oxford; Botnar Research Centre, University of Oxford; Freiburg Institute for Advanced Studies (FRIAS), University of Freiburg) , Bernd Nidetzky (Graz University of Technology; Austrian Centre of Industrial Biotechnology)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Acs Catalysis

State: Published (Approved)
Published: March 2019
Diamond Proposal Number(s): 1221 , 7864 , 9948 , 15433

Abstract: Biosynthesis of 6-deoxy sugars, including L-fucose, involves a mechanistically complex, enzymatic 4,6-dehydration of hex-ose nucleotide precursors as the first committed step. Here, we determined pre- and post-catalytic complex structures of the human GDP-mannose 4,6-dehydratase at atomic resolution. These structures together with results of molecular dynamics simulation and biochemical characterization of wildtype and mutant enzymes reveal elusive mechanistic details of water elimination from GDP-mannose C5’’ and C6’’, coupled to NADP-mediated hydride transfer from C4’’ to C6’’. We show that concerted acid-base catalysis from only two active-site groups, Tyr179 and Glu157, promotes a syn 1,4-elimination from an enol (not an enolate) intermediate. We also show that the overall multistep catalytic reaction involves least position changes of enzyme and substrate groups; and that it proceeds under conserved exploitation of the basic (minimal) catalytic machinery of short-chain dehydrogenase/reductases.

Journal Keywords: Beta-elimination; carbohydrates; enzyme catalysis; reaction mechanism; sugar dehydratase; shortchain dehydrogenase/reductase

Subject Areas: Chemistry, Biology and Bio-materials

Instruments: I02-Macromolecular Crystallography , I03-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography