7PU0

Crystal Structure of Two-Domain Laccase mutant H165A/M199G from Streptomyces griseoflavus

7PU0 の概要

• エントリーDOI: 10.2210/pdb7pu0/pdb
• 分子名称: Two-domain laccase, COPPER (II) ION, SODIUM ION, ... (4 entities in total)
• 機能のキーワード: two-domain laccase, laccase, streptomyces griseoflavus, oxidoreductase
• 由来する生物種: Streptomyces griseoflavus
• タンパク質・核酸の鎖数: 6
• 化学式量合計: 186083.73

構造登録者

Gabdulkhakov, A.,Tishchenko, S.,Kolyadenko, I. (登録日: 2021-09-28, 公開日: 2022-01-19, 最終更新日: 2024-01-31)

主引用文献

Kolyadenko, I.,Scherbakova, A.,Kovalev, K.,Gabdulkhakov, A.,Tishchenko, S.
Engineering the Catalytic Properties of Two-Domain Laccase from Streptomyces griseoflavus Ac-993.
Int J Mol Sci, 23:-, 2021

PubMed Abstract: Laccases catalyze the oxidation of substrates with the concomitant reduction of oxygen to water. Recently, we found that polar residues located in tunnels leading to Cu2 and Cu3 ions control oxygen entrance (His 165) and proton transport (Arg 240) of two-domain laccase (2D) from (SgfSL). In this work, we have focused on optimizing the substrate-binding pocket (SBP) of SgfSL while simultaneously adjusting the oxygen reduction process. SgfSL variants with three single (Met199Ala, Met199Gly, and Tyr230Ala) and three double amino acid residues substitutions (Met199Gly/His165Ala, His165Ala/Arg240His, Met199Gly/Arg240His) were constructed, purified, and investigated. Combination of substitutions in the SBP and in the tunnel leading to Cu2 ion (Met199Gly/Arg240His) increased SgfSL catalytic activity towards ABTS by 5-fold, and towards 2.6-DMP by 16-fold. The high activity of the Met199Gly/Arg240His variant can be explained by the combined effect of the SBP geometry optimization (Met199Gly) and increased proton flux via the tunnel leading to Cu2 ion (Arg240His). Moreover, the variant with Met199Gly and His165Ala mutations did not significantly increase SgfSL's activity, but led to a drastic shift in the optimal pH of 2.6-DMP oxidation. These results indicate that His 165 not only regulates oxygen access, but it also participates in proton transport in 2D laccases.

PubMed: 35008493
DOI: 10.3390/ijms23010065

実験手法

X-RAY DIFFRACTION (2.2 Å)