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The open reading frame 02797 from Candida tropicalis encodes a novel NADH-dependent aldehyde reductase.
Protein Expression and Purification ( IF 1.4 ) Pub Date : 2020-03-12 , DOI: 10.1016/j.pep.2020.105625
Xiangdong Hu 1 , Xuebing Han 1 , Lan Wu 1 , Hanyu Wang 1 , Yidan Ouyang 1 , Qian Li 1 , Xiaolin Kuang 1 , Quanju Xiang 2 , Xiumei Yu 2 , Xi Li 3 , Yunfu Gu 2 , Ke Zhao 2 , Qiang Chen 2 , Menggen Ma 4
Affiliation  

Owing to its high-temperature tolerance, robustness, and wide use of carbon sources, Candida tropicalis is considered a good candidate microorganism for bioconversion of lignocellulose to ethanol. It also has the intrinsic ability to in situ detoxify aldehydes derived from lignocellulosic hydrolysis. However, the aldehyde reductases that catalyze this bioconversion in C. tropicalis remain unknown. Herein, we found that the uncharacterized open reading frame (ORF), CTRG_02797, from C. tropicalis encodes a novel and broad substrate-specificity aldehyde reductase that reduces at least seven aldehydes. This enzyme strictly depended on NADH rather than NADPH as the co-factor for catalyzing the reduction reaction. Its highest affinity (Km), maximum velocity (Vmax), catalytic rate constant (Kcat), and catalytic efficiency (Kcat/Km) were observed when reducing acetaldehyde (AA) and its enzyme activity was influenced by different concentrations of salts, metal ions, and chemical protective additives. Protein localization assay demonstrated that Ctrg_02797p was localized in the cytoplasm in C. tropicalis cells, which ensures an effective enzymatic reaction. Finally, Ctrg_02797p was grouped into the cinnamyl alcohol dehydrogenase (CADH) subfamily of the medium-chain dehydrogenase/reductase family. This research provides guidelines for exploring more uncharacterized genes with reduction activity for detoxifying aldehydes.



中文翻译:

来自热带假丝酵母的开放阅读框02797编码新型的NADH依赖性醛还原酶。

由于其高温耐受性,坚固性和碳源的广泛使用,热带假丝酵母被认为是将木质纤维素生物转化为乙醇的良好候选微生物。它还具有就地解毒源自木质纤维素水解的醛的固有能力。然而,催化这种在热带假丝酵母中的生物转化的醛还原酶仍然未知。在这里,我们发现来自热带假丝酵母的未表征的开放阅读框(ORF)CTRG_02797编码一种新颖且广泛的底物特异性醛还原酶,可还原至少七个醛。该酶严格依赖于NADH而不是NADPH作为催化还原反应的辅助因子。当还原乙醛(AA)时,观察到其最高亲和力(K m),最大速度(V max),催化速率常数(K cat)和催化效率(K cat / K m),并且其酶活性受不同浓度的影响盐,金属离子和化学保护添加剂。蛋白质定位分析表明,Ctrg_02797p位于热带假丝酵母的细胞质中细胞,确保有效的酶促反应。最后,将Ctrg_02797p分组为中链脱氢酶/还原酶家族的肉桂醇脱氢酶(CADH)亚家族。这项研究为探索更多具有还原活性的醛脱毒活性基因提供了指导。

更新日期:2020-03-12
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