Kuang Xiaolin, Ouyang Yidan, Guo Yaping, Li Qian, Wang Hanyu, Abrha Getachew Tafere, Ayepa Ellen, Gu Yunfu, Li Xi, Chen Qiang, Ma Menggen
Institute of Resources and Geographic Information Technology, College of Resources, Sichuan Agricultural University, No. 211 Huimin Road, Wenjiang, Chengdu, 611130, Sichuan, People's Republic of China.
Patent Examination Cooperation Sichuan Center of the Patent Office, SIPO, Chengdu, 610213, Sichuan, People's Republic of China.
Appl Microbiol Biotechnol. 2020 Aug;104(15):6679-6692. doi: 10.1007/s00253-020-10722-9. Epub 2020 Jun 15.
At least 24 aldehyde reductases from Saccharomyces cerevisiae have been characterized and most function in in situ detoxification of lignocellulosic aldehyde inhibitors, but none is classified into the polyol dehydrogenase (PDH) subfamily of the medium-chain dehydrogenase/reductase (MDR) superfamily. This study confirmed that two (2R,3R)-2,3-butanediol dehydrogenases (BDHs) from industrial (denoted Y)/laboratory (denoted B) strains of S. cerevisiae, Bdh1p(Y)/Bdh1p(B) and Bdh2p(Y)/Bdh2p(B), were members of the PDH subfamily with an NAD(P)H binding domain and a catalytic zinc binding domain, and exhibited reductive activities towards lignocellulosic aldehyde inhibitors, such as acetaldehyde, glycolaldehyde, and furfural. Especially, the highest enzyme activity towards acetaldehyde by Bdh2p(Y) was 117.95 U/mg with cofactor nicotinamide adenine dinucleotide reduced (NADH). Based on the comparative kinetic property analysis, Bdh2p(Y)/Bdh2p(B) possessed higher specific activity, substrate affinity, and catalytic efficiency towards glycolaldehyde than Bdh1p(Y)/Bdh1p(B). This was speculated to be related to their 49% sequence differences and five nonsynonymous substitutions (Ser41Thr, Glu173Gln, Ile270Leu, Ile316Met, and Gly317Cys) occurred in their conserved NAD(P)H binding domains. Compared with BDHs from a laboratory strain, Bdh1p(Y) and Bdh2p(Y) from an industrial strain displayed five nonsynonymous mutations (Thr, Asn, Glu, Val, and Ala) and three nonsynonymous mutations (Ala, Ile, and Ala), respectively. From a first analysis with selected aldehydes, their reductase activities were different from BDHs of laboratory strain, and their catalytic efficiency was higher towards glycolaldehyde and lower towards acetaldehyde. Comparative investigation of kinetic properties of BDHs from S. cerevisiae as aldehyde reductases provides a guideline for their practical applications in in situ detoxification of aldehyde inhibitors during lignocellulose bioconversion.Key Points• Two yeast BDHs have enzyme activities for reduction of aldehydes.• Overexpression of BDHs slightly improves yeast tolerance to acetaldehyde and glycolaldehyde.• Bdh1p and Bdh2p differ in enzyme kinetic properties.• BDHs from strains with different genetic backgrounds differ in enzyme kinetic properties.
酿酒酵母中至少有24种醛还原酶已被鉴定，大多数在木质纤维素醛抑制剂的原位解毒中发挥作用，但没有一种被归类到中链脱氢酶/还原酶（MDR）超家族的多元醇脱氢酶（PDH）亚家族中。本研究证实，来自酿酒酵母工业菌株（记为Y）/实验室菌株（记为B）的两种（2R,3R）-2,3-丁二醇脱氢酶（BDH），即Bdh1p(Y)/Bdh1p(B)和Bdh2p(Y)/Bdh2p(B)，是具有NAD(P)H结合结构域和催化锌结合结构域的PDH亚家族成员，并对木质纤维素醛抑制剂，如乙醛、乙醇醛和糠醛，表现出还原活性。特别是，Bdh2p(Y)对乙醛的最高酶活性在辅酶烟酰胺腺嘌呤二核苷酸还原型（NADH）存在下为117.95 U/mg。基于比较动力学性质分析，Bdh2p(Y)/Bdh2p(B)对乙醇醛的比活性、底物亲和力和催化效率高于Bdh1p(Y)/Bdh1p(B)。据推测，这与它们49%的序列差异以及在其保守的NAD(P)H结合结构域中发生的五个非同义替换（Ser41Thr、Glu173Gln、Ile270Leu,、Ile316Met和Gly317Cys）有关。与来自实验室菌株的BDH相比，来自工业菌株的Bdh1p(Y)和Bdh2p(Y)分别显示出五个非同义突变（Thr,、Asn、Glu、Val和Ala）和三个非同义突变（Ala、Ile和Ala）。从对选定醛类的初步分析来看，它们的还原酶活性与实验室菌株的BDH不同，且它们对乙醇醛的催化效率较高，对乙醛的催化效率较低。对酿酒酵母BDH作为醛还原酶的动力学性质进行比较研究，为它们在木质纤维素生物转化过程中醛抑制剂的原位解毒中的实际应用提供了指导。要点：• 两种酵母BDH具有还原醛类的酶活性。• BDH的过表达略微提高了酵母对乙醛和乙醇醛的耐受性。• Bdh1p和Bdh2p在酶动力学性质上存在差异。• 来自不同遗传背景菌株的BDH在酶动力学性质上存在差异。
