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Substrate-imprinted docking of Agrobacterium tumefaciens uronate dehydrogenase for increased substrate selectivity.

Abstract：

:Agrobacterium tumefaciens uronate dehydrogenase (AtuUdh) belongs to the short-chain dehydrogenase superfamily, specifically those acting on the CH-OH group of donor with NAD+ or NADP+ as acceptor. It is apparently required for the production of D-glucaric acid. AtuUdh-catalyzed reaction is reversible with dual substrate-specific activity (D-galacturonic acid and D-glucuronic acid) in nature. In our study, 34 mutants were pre-screened from 155 mutants generated from AtuUdh (wild-type) and selected 10 structurally stable mutants with increased substrate selectivity. The specificity, efficiency, and selectivity of these mutants for different substrates and cofactors were predicted from 121 docked models using a substrate-imprinted docking approach. Q14F, S36L, and S75T mutants have shown a high binding affinity to D-glucuronic acid and its substrate intermediates such as D-glucaro-1,4-lactone and D-glucaro-1,5-lactone. These mutants exhibited a low binding affinity to the substrate and cofactor required for D-galactaric acid. D34S, N112E and S165E mutants found to show a high selectivity of D-galacturonic acid and its substrate intermediates for D-galactaric acid production. Ser75, Ser165, and Arg174 are active residues playing an imperative role in the substrate selectivity and also contributed in the conjecture the mechanism of transition state stabilization catalyzed by AtuUdh mutants. The present approach was used to reveal the substrate binding mechanism of AtuUdh mutants for a better understanding of the structural basis for selectivity and function.

journal_name

Int J Biol Macromol

journal_title

International journal of biological macromolecules

authors

Murugan A,Prathiviraj R,Mothay D,Chellapandi P

doi

10.1016/j.ijbiomac.2019.08.194

subject

Has Abstract

pub_date

2019-11-01 00:00:00

pages

1214-1225

eissn

0141-8130

issn

1879-0003

pii

S0141-8130(19)33393-8

journal_volume

140

pub_type

杂志文章

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