title: Unusual carboxylesterase bearing a GGG(A)X-type oxyanion hole discovered in Paenibacillus barcinonensis BP-23.
abstract: Strain Paenibacillus barcinonensis BP-23, previously isolated from Ebro's river delta (Spain), bears a complex hydrolytic system showing the presence of at least two enzymes with activity on lipidic substrates. EstA, a cell-bound B-type carboxylesterase from the strain was previously isolated and characterized. The gene coding for a second putative lipase, located upstream cellulase Cel5A, was obtained using a genome walking strategy and cloned in Escherichia coli for further characterization. The recombinant clone obtained displayed high activity on medium/short-chain fatty acid-derivative substrates. The enzyme, named Est23, was purified and characterized, showing maximum activity on pNP-caprylate (C8:0) or MUF-heptanoate (C7:0) under conditions of moderate temperature and pH. Although Est23 displays a GGG(A)X-type oxyanion hole, described as an important motif for tertiary alcohol ester resolution, neither conversion nor enantiomeric resolution of tertiary alcohols could be detected. Amino acid sequence alignment of Est23 with those of known bacterial lipase families and with closely related proteins suggests that the cloned enzyme does not belong to any of the described bacterial lipase families. A phylogenetic tree including Est23 and similar amino acid sequences showed that the enzyme belongs to a differentiated sequence cluster which probably constitutes a new family of bacterial lipolytic enzymes.
authors.affiliation: Department of Microbiology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain.
authors.forename: Belén
authors.initials: B
authors.lastname: Infanzón
authors.affiliation: Department of Microbiology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain.
authors.forename: Susana V
authors.initials: SV
authors.lastname: Valenzuela
authors.affiliation: Department of Microbiology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain.
authors.forename: Amanda
authors.initials: A
authors.lastname: Fillat
authors.affiliation: Department of Microbiology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain.
authors.forename: F I Javier
authors.initials: FI
authors.lastname: Pastor
authors.affiliation: Department of Microbiology, University of Barcelona, Av. Diagonal 643, 08028 Barcelona, Spain. Electronic address: pdiaz@ub.edu.
authors.forename: Pilar
authors.initials: P
authors.lastname: Diaz
mesh_terms: D000438:Alcohols / Q000378:metabolism; D020816:Amino Acid Motifs; D043182:Carboxylesterase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D017124:Conserved Sequence; D056507:Paenibacillus / Q000201:enzymology*; D010447:Peptide Hydrolases / Q000378:metabolism; D010802:Phylogeny
chemical_list: D000438:Alcohols; D043182:Carboxylesterase; D010447:Peptide Hydrolases
keywords: Carboxylesterases; Paenibacillus