title: Isolation and characterization of a family VII esterase derived from alluvial soil metagenomic library.
abstract: A novel esterase gene, estDL30, was isolated from an alluvial metagenomic library using function-driven screening. estDL30 consisted of 1,524 nucleotides and encoded a 507-amino acid protein. Sequence analysis revealed that EstDL30 is similar to many type B carboxylesterases, containing a G-E-S-A-G pentapeptide with a catalytic Ser residue. Phylogenetic analysis suggested that EstDL30 belongs to the family VII lipases, together with esterases from Bacillus subtilis (P37967), Streptomyces coelicolor A3(2) (CAA22794), and Arthrobacter oxydans (Q01470). Purified EstDL30 showed its highest catalytic efficiency toward p-nitrophenyl butyrate, with a k (cat) of 2293 s(-1) and k (cat)/K (m) of 176.4 s(-1)mM(-1); however, little activity was detected when the acyl chain length exceeded C(8). Biochemical characterization of EstDL30 revealed that it is an alkaline esterase that possesses maximal activity at pH 8 and 40° C. The effects of denaturants and divalent cations were also investigated. EstDL30 tolerated well the presence of methanol and Tween 20. Its activity was strongly inhibited by 1 mM Cu(2+) and Zn(2+), but stimulated by Fe(2+). The unique properties of EstDL30, its high activity under alkaline conditions and stability in the presence of organic solvents, may render it applicable to organic synthesis.
authors.affiliation: Department of Medical Bioscience, Dong-A University, Busan 604-714, Republic of Korea.
authors.forename: Weixin
authors.initials: W
authors.lastname: Tao
authors.forename: Myung Hwan
authors.initials: MH
authors.lastname: Lee
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.forename: Nam Hee
authors.initials: NH
authors.lastname: Kim
authors.forename: Seon-Woo
authors.initials: SW
authors.lastname: Lee
mesh_terms: D020816:Amino Acid Motifs; D001173:Arthrobacter / Q000201:enzymology / Q000235:genetics; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D002087:Butyrates / Q000378:metabolism; D020134:Catalytic Domain; D016000:Cluster Analysis; D003067:Coenzymes / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D004950:Esterases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D054892:Metagenome; D008969:Molecular Sequence Data; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012988:Soil Microbiology; D048372:Streptomyces coelicolor / Q000201:enzymology / Q000235:genetics; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D002087:Butyrates; D003067:Coenzymes; D004791:Enzyme Inhibitors; C033592:4-nitrophenyl butyrate; D004950:Esterases