Implications for an imidazol-2-yl carbene intermediate in the rhodanase-catalyzed C-S bond formation reaction of anaerobic ergothioneine biosynthesis.
Implications for an imidazol-2-yl carbene intermediate in the rhodanase-catalyzed C-S bond formation reaction of anaerobic ergothioneine biosynthesis.
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DOI：
10.1021/acscatal.0c04886
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发表时间：
2021-03-19
期刊：
影响因子：
12.9
通讯作者：
Liu P
中科院分区：
文献类型：
--
作者：
Cheng R;Lai R;Peng C;Lopez J;Li Z;Naowarojna N;Li K;Wong C;Lee N;Whelan SA;Qiao L;Grinstaff MW;Wang J;Cui Q;Liu P
在厌氧麦角硫因生物合成途径中，含有罗丹酸结构域的酶（EanB）激活组氨酸的sp2 ε-C-H Dona，并用C-S键取代它以产生麦角硫因。该转硫化反应的关键中间体是Cys 412过硫化物。用Cys 412全硒化物取代EanB-Cys 412全硫化物不产生麦角硫因的硒类似物硒因。而在氘代缓冲液中，过硒化修饰的EanB催化组氨酸的sp2 ε-C-H键与D2 O之间的氘交换。从QM/MM计算结果表明，该反应涉及的卡宾中间体和Tyr 353起着关键作用。我们假设调节Tyr 353的pKa将影响氘交换速率。事实上，含有3，5-二氟酪氨酸的EanB催化氘交换反应，kex比野生型EanB（EanBWT）高约10倍。关于潜在的机制，这些结果支持参与的卡宾中间体在EanB-催化，使EanB作为为数不多的卡宾中间体涉及酶系统之一。
In the anaerobic ergothioneine biosynthetic pathway, a rhodanese domain containing enzyme (EanB) activates tne hercynine’s sp2 ε-C-H Dona ana replaces it with a C-S bond to produce ergothioneine. The key intermediate for this trans-sulfuration reaction is the Cys412 persulfide. Substitution of the EanB-Cys412 persulfide with a Cys412 perselenide does not yield the selenium analog of ergothioneine, selenoneine. However, in deuterated buffer, the perselenide-modified EanB catalyzes the deuterium exchange between hercynine’s sp2 ε-C-H bond and D2O. Results from QM/MM calculations suggest that the reaction involves a carbene intermediate and that Tyr353 plays a key role. We hypothesize that modulating the pKa of Tyr353 will affect the deuterium-exchange rate. Indeed, the 3,5-difluoro tyrosine containing EanB catalyzes the deuterium exchange reaction with kex of ~10-fold greater than the wild-type EanB (EanBWT). With regards to potential mechanisms, these results support the involvement of a carbene intermediate in EanB-catalysis, rendering EanB as one of the few carbene-intermediate involving enzymatic systems.
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