title: Novel thermophilic and thermostable lipolytic enzymes from a Thailand hot spring metagenomic library.
abstract: Functional screening for lipolytic enzymes from a metagenomic library (origin: Jae Sawn hot spring, Thailand) resulted in isolation of a novel patatin-like phospholipase (PLP) and an esterase (Est1). PLP contained four conserved domains similar to other patatin-like proteins with lipid acyl hydrolase activity. Likewise, sequence alignment analysis revealed that Est1 can be classified as a family V bacterial lipolytic enzyme. Both PLP and Est1 were expressed heterologously as soluble proteins in E. coli and exhibited more than 50% of their maximal activities at alkaline pH, of 7-9 and 8-10, respectively. In addition, both enzymes retained more than 50% of maximal activity in the temperature range of 50-75 degrees C, with optimal activity at 70 degrees C and were stable at 70 degrees C for at least 120 min. Both PLP and Est1 exhibited high V(max) toward p-nitrophenyl butyrate. The enzymes had activity toward both short-chain (C(4) and C(5)) and long chain (C(14) and C(16)) fatty acid esters. The isolated enzymes, are therefore, different from other known patatin-like phospholipases and esterases, which usually show no activity for substrates longer than C(10). We suggest that PLP and EstA enzymes are novel and have a; b potential use in industrial applications.
authors.affiliation: BIOTEC Central Research Unit, National Center for Genetic Engineering and Biotechnology, 113 National Science Park, Paholyothin Road, Klong 1, Klong Luang, Pathumthani 12120, Thailand.
authors.forename: Pacawadee
authors.initials: P
authors.lastname: Tirawongsaroj
authors.forename: Rutchadaporn
authors.initials: R
authors.lastname: Sriprang
authors.forename: Piyanun
authors.initials: P
authors.lastname: Harnpicharnchai
authors.forename: Taksawan
authors.initials: T
authors.lastname: Thongaram
authors.forename: Verawat
authors.initials: V
authors.lastname: Champreda
authors.forename: Sutipa
authors.initials: S
authors.lastname: Tanapongpipat
authors.forename: Kusol
authors.initials: K
authors.lastname: Pootanakit
authors.forename: Lily
authors.initials: L
authors.lastname: Eurwilaichitr
mesh_terms: D001483:Base Sequence; D002874:Chromosome Mapping; D004789:Enzyme Activation; D004795:Enzyme Stability; D004950:Esterases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D045482:Hot Springs / Q000382:microbiology*; D008055:Lipids / Q000737:chemistry; D008066:Lipolysis; D008969:Molecular Sequence Data; D019151:Peptide Library; D010740:Phospholipases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D013045:Species Specificity; D013696:Temperature; D013785:Thailand
chemical_list: D008055:Lipids; D019151:Peptide Library; D004950:Esterases; D010740:Phospholipases