A Novel Esterase from Paenibacillus sp. PBS-2 Is a New Member of the ��-Lactamase Belonging to the Family VIII Lipases/Esterases

Abstract

Screening of a gene library from Paenibacillus sp. PBS-2 generated in Escherichia coli led to the identification of a clone with lipolytic activity. Sequence analysis showed an open reading frame encoding a polypeptide of 378 amino acid residues with a predicted molecular mass of 42 kDa. The esterase displayed 69% and 42% identity with the putative β-lactamases from Paenibacillus sp. JDR-2 and Clostridium sp. BNL1100, respectively. The esterase contained a Serx-x-Lys motif that is conserved among all β-lactamases found to date. The protein PBS-2 was produced in both soluble and insoluble forms when E. coli cells harboring the gene were cultured at 18°C. The enzyme is a serine protein and was active against p-nitrophenyl esters of C 2 , C 4 , C 8 , and C 10 . The optimum pH and temperature for enzyme activity were pH 9.0 and 30°C, respectively. Relative activity of 55% remained at up to 5°C with an activation energy of 5.84 kcal/mol, which indicates that the enzyme is cold-adapted. Enzyme activity was inhibited by Cd 2+ , Cu 2+ , and Hg 2+ ions. As expected for a serine esterase, activity was inhibited by phenylmethylsulfonyl fluoride. The enzyme was remarkably active and stable in the presence of commercial detergents and organic solvents. This cold-adapted esterase has potential as a biocatalyst and detergent additive for use at low temperatures.

Molecular Characterization of a Novel Family VIII Esterase with β-Lactamase Activity (PsEstA) from Paenibacillus sp.

“…Cloning and expression of PsEstA were performed as previously described [18]. Transformed E. coli BL21(λDE3) cells were grown in LB medium until OD 600 reached 0.4-0.6.…”

“…A novel family VIII esterase with β-lactamase activity (PsEstA) has recently been identified in Paenibacillus sp. PBS-2 [18]. Here, we biochemically characterized PsEstA to lay the foundation for the understanding of this enzyme at the molecular level.…”

Biochemical and Structural Analysis of a Novel Esterase from Caulobacter crescentus related to Penicillin-Binding Protein (PBP)

“…PBS-2, a bacterial PBP homologue from Paenibacillus sp. , was used as a positive control55. Note that CcEstA showed almost no hydrolytic activity toward 7-ACA and CTX, while nitrocefin can be hydrolyzed.…”

Cloning, expression, and biochemical characterization of a cold-active GDSL-esterase of a Pseudomonas sp. S9 isolated from Spitsbergen island soil.

“…strain S9. In contrast to most well characterized cold-active esterases containing a conserved G-X-S-X-G consensus sequence centered around the catalytic serine (Suzuki et al, 2002;Kulakova et al, 2004;Soror et al, 2007;Heath et al, 2009;Fu et al, 2011;Kang et al, 2011;Brault et al, 2012;Hu et al, 2012;Jiang et al, 2012;Jimenez et al, 2012;Novototskaya-Vlasova et al, 2012;Lemak et al, 2012;Abdul Salam et al, 2013;Berlemont et al, 2013;Fu et al, 2013;Kim et al, 2014) the EstS9 harbors another conserved sequence G-D-S-L around this catalytic residue. To the best of our knowledge, the EstS9 is the third cold-active esterase belonging to the GDSL subfamily of lipolytic enzymes (Suzuki et al, 2003;Cieslinski et al, 2007) described to date.…”