Li Ping-Yi, Yao Qiong-Qiong, Wang Peng, Zhang Yi, Li Yue, Zhang Yan-Qi, Hao Jie, Zhou Bai-Cheng, Chen Xiu-Lan, Shi Mei, Zhang Yu-Zhong, Zhang Xi-Ying
State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China
Appl Environ Microbiol. 2017 Apr 17;83(9). doi: 10.1128/AEM.00131-17. Print 2017 May 1.
Microbial esterases play important roles in deep-sea organic carbon degradation and cycling. Although they have similar catalytic triads and oxyanion holes, esterases are hydrolases and homoserine transacetylases (HTAs) are transferases. Because two HTA homologs were identified as acetyl esterases, the HTA family was recently divided into the acetyltransferase subfamily and the acetyl esterase subfamily. Here, we identified and characterized a novel HTA-like esterase, Est22, from a deep-sea sedimentary metagenomic library. Est22 could efficiently hydrolyze esters with acyl lengths of up to six carbon atoms but had no transacetylase activity, which is different from HTAs and HTA-like acetyl esterases. Phylogenetic analysis also showed that Est22 and its homologs form a separate branch of the HTA family. We solved the structures of Est22 and its L374D mutant and modeled the structure of the L374D mutant with -nitrophenyl butyrate. Based on structural, mutational, and biochemical analyses, Phe and Met in the oxyanion hole and Arg were revealed to be the key substrate-binding residues. A detailed structural comparison indicated that differences in their catalytic tunnels lead to the different substrate specificities of Est22 and the other two HTA subfamilies. Biochemical and sequence analyses suggested that Est22 homologs may have the same substrate recognition and catalysis mechanisms as Est22. Due to the significant differences in sequences, structures, and substrate specificities between Est22 (and its homologs) and the other two HTA subfamilies, we suggest that Est22 and its homologs represent a new subfamily in the HTA family. Microbial esterases play important roles in the turnover of organic carbon in the deep sea. Esterases and HTAs represent two groups of α/β hydrolases. Esterases catalyze the hydrolysis of simple esters and are widely used in the pharmaceutical and agrochemical industries, while HTAs catalyze the transfer of an acetyl group from acetyl-coenzyme A (CoA) to homoserine and are essential for microbial growth. Here, we report on a novel HTA-like esterase, Est22, from a deep-sea sediment. Because of the significant differences in sequences, structures, and substrate specificities of HTAs and HTA-like acetyl esterases, Est22 and its homologs represent a new subfamily in the HTA family. This study offers new knowledge regarding marine esterases.
微生物酯酶在深海有机碳降解和循环中发挥着重要作用。尽管它们具有相似的催化三联体和氧阴离子洞，但酯酶是水解酶，而高丝氨酸转乙酰酶（HTAs）是转移酶。由于两种HTA同源物被鉴定为乙酰酯酶，HTA家族最近被分为乙酰转移酶亚家族和乙酰酯酶亚家族。在此，我们从一个深海沉积宏基因组文库中鉴定并表征了一种新型的HTA样酯酶Est22。Est22能够高效水解酰基长度达六个碳原子的酯，但没有转乙酰酶活性，这与HTAs和HTA样乙酰酯酶不同。系统发育分析还表明，Est22及其同源物在HTA家族中形成一个单独的分支。我们解析了Est22及其L374D突变体的结构，并用丁酸对硝基苯酯对L374D突变体的结构进行了建模。基于结构、突变和生化分析，揭示了氧阴离子洞中的苯丙氨酸和甲硫氨酸以及精氨酸是关键的底物结合残基。详细的结构比较表明，它们催化通道的差异导致了Est22与其他两个HTA亚家族底物特异性的不同。生化和序列分析表明，Est22同源物可能具有与Est22相同的底物识别和催化机制。由于Est22（及其同源物）与其他两个HTA亚家族在序列、结构和底物特异性上存在显著差异，我们认为Est22及其同源物代表了HTA家族中的一个新亚家族。微生物酯酶在深海有机碳周转中发挥着重要作用。酯酶和HTAs代表两组α/β水解酶。酯酶催化简单酯的水解，广泛应用于制药和农用化学工业，而HTAs催化乙酰基从乙酰辅酶A（CoA）转移至高丝氨酸，对微生物生长至关重要。在此，我们报道了一种来自深海沉积物的新型HTA样酯酶Est22。由于HTAs和HTA样乙酰酯酶在序列、结构和底物特异性上存在显著差异，Est22及其同源物代表了HTA家族中的一个新亚家族。这项研究为海洋酯酶提供了新的知识。
