title: A Novel Subfamily Esterase with a Homoserine Transacetylase-like Fold but No Transferase Activity.
abstract: 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 thebona fideacetyltransferase 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 withp-nitrophenyl butyrate. Based on structural, mutational, and biochemical analyses, Phe71and Met176in the oxyanion hole and Arg294were 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.IMPORTANCEMicrobial 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.
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Ping-Yi
authors.initials: PY
authors.lastname: Li
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Qiong-Qiong
authors.initials: QQ
authors.lastname: Yao
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Li
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Yan-Qi
authors.initials: YQ
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Hao
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Bai-Cheng
authors.initials: BC
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China cxl0423@sdu.edu.cn zhangxiying@sdu.edu.cn.
authors.forename: Xiu-Lan
authors.initials: XL
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.
authors.forename: Mei
authors.initials: M
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China.|Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao, China.
authors.forename: Yu-Zhong
authors.initials: YZ
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Technology, Marine Biotechnology Research Center, Institute of Marine Science and Technology, Shandong University, Jinan, China cxl0423@sdu.edu.cn zhangxiying@sdu.edu.cn.
authors.forename: Xi-Ying
authors.initials: XY
authors.lastname: Zhang
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D016000:Cluster Analysis; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D004950:Esterases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004952:Esters / Q000378:metabolism; D006868:Hydrolysis; D054892:Metagenome; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011487:Protein Conformation; D012623:Seawater; D017385:Sequence Homology
chemical_list: D004952:Esters; D000123:Acetyltransferases; C021369:homoserine O-acetyltransferase; D004950:Esterases
keywords: catalysis; crystal structure; new HTA subfamily; serine esterase; substrate recognition