脱水酶
化学
二价
活动站点
裂解酶
结合位点
立体化学
QM/毫米
对接(动物)
组合化学
催化作用
酶
生物化学
有机化学
医学
护理部
作者
Tenuun Bayaraa,Thierry Lonhienne,Samuel Sutiono,Okke Melse,Thomas Brück,Esteban Marcellin,Paul V. Bernhardt,Mikael Bodén,Jeffrey R. Harmer,Volker Sieber,Luke W. Guddat,Gerhard Schenk
标识
DOI:10.1002/chem.202203140
摘要
Enzyme-catalyzed reaction cascades play an increasingly important role for the sustainable manufacture of diverse chemicals from renewable feedstocks. For instance, dehydratases from the ilvD/EDD superfamily have been embedded into a cascade to convert glucose via pyruvate to isobutanol, a platform chemical for the production of aviation fuels and other valuable materials. These dehydratases depend on the presence of both a Fe-S cluster and a divalent metal ion for their function. However, they also represent the rate-limiting step in the cascade. Here, catalytic parameters and the crystal structure of the dehydratase from Paralcaligenes ureilyticus (PuDHT, both in presence of Mg2+ and Mn2+ ) were investigated. Rate measurements demonstrate that the presence of stoichiometric concentrations Mn2+ promotes higher activity than Mg2+ , but at high concentrations the former inhibits the activity of PuDHT. Molecular dynamics simulations identify the position of a second binding site for the divalent metal ion. Only binding of Mn2+ (not Mg2+ ) to this site affects the ligand environment of the catalytically essential divalent metal binding site, thus providing insight into an inhibitory mechanism of Mn2+ at higher concentrations. Furthermore, in silico docking identified residues that play a role in determining substrate binding and selectivity. The combined data inform engineering approaches to design an optimal dehydratase for the cascade.
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