3,6-Anhydro-L-Galactose Dehydrogenase VvAHGD is a Member of a New Aldehyde Dehydrogenase Family and Catalyzes by a Novel Mechanism with Conformational Switch of Two Catalytic Residues Cysteine 282 and Glutamate 248
3,6-Anhydro-L-Galactose Dehydrogenase VvAHGD is a Member of a New Aldehyde Dehydrogenase Family and Catalyzes by a Novel Mechanism with Conformational Switch of Two Catalytic Residues Cysteine 282 and Glutamate 248
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3,6-脱水-L-半乳糖脱氢酶 VvAHGD 是新醛脱氢酶家族的成员，通过具有两个催化残基半胱氨酸 282 和谷氨酸 248 构象转换的新颖机制进行催化
DOI：
10.1016/j.jmb.2020.02.008
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发表时间：
2020-03-27
影响因子：
5.6
通讯作者：
Zhang, Yu-Zhong
中科院分区：
文献类型：
--
作者：
Wang, Yue;Li, Ping-Yi;Zhang, Yu-Zhong
3,6-无水α - l -半乳糖（L-AHG）是红藻的主要单糖成分之一。在最近发现的细菌L-AHG分解代谢途径中，L-AHG首先被NAD(P)（+）依赖性脱氢酶（AHGD）氧化，这是该途径的关键步骤。然而，AHGDs的催化机制尚不清楚。在这里，我们鉴定并鉴定了来自海洋细菌可变弧菌JCM 19239 （VvAHGD）的AHGD。NADP（+）依赖性的VvAHGD能有效氧化L-AHG。系统发育分析表明，VvAHGD及其同源物代表一个新的醛脱氢酶（ALDH）家族，与已有报道的ALDH家族具有不同的底物偏好，被命名为L-AHGDH家族。为了解释VvAHGD的催化机理，我们求解了VvAHGD以载脂蛋白形式和NADP（+）络合物的结构，并用L-AHG对其结构进行了建模。基于结构、突变和生化分析，确定了VvAHGD的辅因子通道和底物通道，揭示了参与NADP（+）和L-AHG结合及其催化作用的关键残基。VvAHGD通过其两个催化残基Cys282和Glu248的构象变化，控制辅因子通道和底物通道的连续连接和中断，从而实现催化作用。结构和酶动力学的比较分析表明，底物通道（形状、大小、静电表面和残留物组成）的差异导致VvAHGD与其他aldh对底物的偏好不同。这项研究揭示了NAD(P)（+）依赖性aldh的多种催化机制和演化。(C) 2020 Elsevier Ltd.版权所有。
3,6-anhydro-alpha-L-galactose (L-AHG) is one of the main monosaccharide constituents of red macroalgae. In the recently discovered bacterial L-AHG catabolic pathway, L-AHG is first oxidized by a NAD(P)(+)-dependent dehydrogenase (AHGD), which is a key step of this pathway. However, the catalytic mechanism(s) of AHGDs is still unclear. Here, we identified and characterized an AHGD from marine bacterium Vibrio variabilis JCM 19239 (VvAHGD). The NADP(+)-dependent VvAHGD could efficiently oxidize L-AHG. Phylogenetic analysis suggested that VvAHGD and its homologs represent a new aldehyde dehydrogenase (ALDH) family with different substrate preferences from reported ALDH families, named the L-AHGDH family. To explain the catalytic mechanism of VvAHGD, we solved the structures of VvAHGD in the apo form and complex with NADP(+) and modeled its structure with L-AHG. Based on structural, mutational, and biochemical analyses, the cofactor channel and the substrate channel of VvAHGD are identified, and the key residues involved in the binding of NADP(+) and L-AHG and the catalysis are revealed. VvAHGD performs catalysis by controlling the consecutive connection and interruption of the cofactor channel and the substrate channel via the conformational changes of its two catalytic residues Cys282 and Glu248. Comparative analyses of structures and enzyme kinetics revealed that differences in the substrate channels (in shape, size, electrostatic surface, and residue composition) lead to the different substrate preferences of VvAHGD from other ALDHs. This study on VvAHGD sheds light on the diversified catalytic mechanisms and evolution of NAD(P)(+)-dependent ALDHs. (C) 2020 Elsevier Ltd. All rights reserved.