Faculty of Biology - Microbiology, Albert-Ludwigs-Universität Freiburg, Schänzlestrasse 1, 79104, Freiburg, Germany.
Max-Planck-Institute for Biophysics, Max-von-Laue-Strasse 3, 60438, Frankfurt, Germany.
Chembiochem. 2021 Nov 16;22(22):3173-3177. doi: 10.1002/cbic.202100421. Epub 2021 Sep 30.
The biologically important, FAD-containing acyl-coenzyme A (CoA) dehydrogenases (ACAD) usually catalyze the anti-1,2-elimination of a proton and a hydride of aliphatic CoA thioesters. Here, we report on the structure and function of an ACAD from anaerobic bacteria catalyzing the unprecedented 1,4-elimination at C3 and C6 of cyclohex-1-ene-1-carboxyl-CoA (Ch1CoA) to cyclohex-1,5-diene-1-carboxyl-CoA (Ch1,5CoA) and at C3 and C4 of the latter to benzoyl-CoA. Based on high-resolution Ch1CoA dehydrogenase crystal structures, the unorthodox reactivity is explained by the presence of a catalytic aspartate base (D91) at C3, and by eliminating the catalytic glutamate base at C1. Moreover, C6 of Ch1CoA and C4 of Ch1,5CoA are positioned towards FAD-N5 to favor the biologically relevant C3,C6- over the C3,C4-dehydrogenation activity. The C1,C2-dehydrogenation activity was regained by structure-inspired amino acid exchanges. The results provide the structural rationale for the extended catalytic repertoire of ACADs and offer previously unknown biocatalytic options for the synthesis of cyclic 1,3-diene building blocks.
生物重要的含 FAD 的酰基辅酶 A (CoA) 脱氢酶（ACAD）通常催化脂肪族 CoA 硫酯的反式 1,2-消除质子和氢化物。在这里，我们报告了一种来自厌氧细菌的 ACAD 的结构和功能，它催化前所未有的 1,4-消除，在环己-1-烯-1-羧基-CoA（Ch1CoA）的 C3 和 C6 处生成环己-1,5-二烯-1-羧基-CoA（Ch1,5CoA），在后者的 C3 和 C4 处生成苯甲酰-CoA。基于高分辨率的 Ch1CoA 脱氢酶晶体结构，通过在 C3 处存在催化天冬氨酸碱（D91），以及消除 C1 处的催化谷氨酸碱，解释了这种非传统的反应性。此外，Ch1CoA 的 C6 和 Ch1,5CoA 的 C4 朝向 FAD-N5 定位，有利于生物相关的 C3,C6-脱氢活性，而不是 C3,C4-脱氢活性。通过结构启发的氨基酸交换恢复了 C1,C2-脱氢活性。这些结果为 ACAD 的扩展催化谱提供了结构基础，并为合成环状 1,3-二烯构建块提供了以前未知的生物催化选择。
