Structures of the Apo and FAD-bound forms of 2-hydroxybiphenyl 3-monooxygenase (HbpA) locate activity hotspots identified by using directed evolution

Chembiochem : a European Journal of Chemical Biology
Chantel N JensenGideon Grogan

Abstract

The FAD-dependent monooxygenase HbpA from Pseudomonas azelaica HBP1 catalyses the hydroxylation of 2-hydroxybiphenyl (2HBP) to 2,3-dihydroxybiphenyl (23DHBP). HbpA has been used extensively as a model for studying flavoprotein hydroxylases under process conditions, and has also been subjected to directed-evolution experiments that altered its catalytic properties. The structure of HbpA has been determined in its apo and FAD-complex forms to resolutions of 2.76 and 2.03 Å, respectively. Comparisons of the HbpA structure with those of homologues, in conjunction with a model of the reaction product in the active site, reveal His48 as the most likely acid/base residue to be involved in the hydroxylation mechanism. Mutation of His48 to Ala resulted in an inactive enzyme. The structures of HbpA also provide evidence that mutants achieved by directed evolution that altered activity are comparatively remote from the substrate-binding site.

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Citations

Mar 25, 2016·Bioscience, Biotechnology, and Biochemistry·Yuan GuoDefu Chen
Jan 4, 2018·Chembiochem : a European Journal of Chemical Biology·Almog Bregman-CohenAyelet Fishman
Jul 10, 2019·Acta Crystallographica. Section F, Structural Biology Communications·John T LazarGeorge Minasov

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