title: A new esterase, belonging to hormone-sensitive lipase family, cloned from Rheinheimera sp. isolated from industrial effluent.
abstract: The gene for esterase (rEst1) was isolated from a new species of genus Rheinheimera by functional screening of E. coli cells transformed with the pSMART/HaeIII genomic library. E. coli cells harboring the esterase gene insert could grow and produce clear halo zones on tributyrin agar. The rEst1 ORF consisted of 1,029 bp, corresponding to 342 amino acid residues with a molecular mass of 37 kDa. The signal P program 3.0 revealed the presence of a signal peptide of 25 amino acids. Esterase activity, however, was associated with a homotrimeric form of molecular mass 95 kDa and not with the monomeric form. The deduced amino acid sequence showed only 54% sequence identity with the closest lipase from Cellvibrio japonicus strain Ueda 107. Conserved domain search and multiple sequence alignment revealed the presence of an esterase/ lipase conserved domain consisting of a GXSXG motif, HGGG motif (oxyanion hole) and HGF motif, typical of the class IV hormone sensitive lipase family. On the basis of the sequence comparison with known esterases/ lipases, REst1 represents a new esterase belonging to class IV family. The purified enzyme worked optimally at 50 degrees C and pH 8, utilized pNP esters of short chain lengths, and showed best catalytic activity with p-nitrophenyl butyrate (C₄), indicating that it was an esterase. The enzyme was completely inhibited by PMSF and DEPC and showed moderate organotolerance.
authors.affiliation: Departments of Microbiology, Panjab University, Chandigarh 160014, India.
authors.forename: Antar Puneet
authors.initials: AP
authors.lastname: Virk
authors.forename: Prince
authors.initials: P
authors.lastname: Sharma
authors.forename: Neena
authors.initials: N
authors.lastname: Capalash
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D002841:Chromatiaceae / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D003470:Culture Media / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015698:Genomic Library; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D055503:Protein Multimerization; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D002787:Sterol Esterase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature; D014169:Transformation, Bacterial; D014280:Triglycerides / Q000378:metabolism
chemical_list: D003470:Culture Media; D004791:Enzyme Inhibitors; D021382:Protein Sorting Signals; D014280:Triglycerides; D002787:Sterol Esterase; C005830:tributyrin