Isolation and characterization of cold-active family VIII esterases from an arctic soil metagenome

• Published in: Applied microbiology and biotechnology 2011-04, Vol.90 (2), p.573-581
• Main Authors: Yu, Eun Young, Kwon, Min-A, Lee, Miae, Oh, Joon Young, Choi, Ji-Eun, Lee, Ji Young, Song, Bong-Keun, Hahm, Dae-Hyun, Song, Jae Kwang
• Format: Article
• Language: English
• Subjects: Amino acids; Arctic Regions; Bacillus cereus; Bacterial Proteins - genetics; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; beta-Lactamases - metabolism; Biochemical Phenomena; Biological and medical sciences; Biomedical and Life Sciences; Biotechnologically Relevant Enzymes and Proteins; Biotechnology; Catalysts; Cloning; Cloning, Molecular; Cold; Cold Temperature; E coli; Enzymes; Escherichia coli; Escherichia coli - genetics; Escherichia coli - metabolism; Esterases - genetics; Esterases - isolation & purification; Esterases - metabolism; Esters; Fundamental and applied biological sciences. Psychology; Gene amplification; Genomes; Genomic Library; Genomics; Hydrogen-Ion Concentration; Life Sciences; Lipase - genetics; Lipase - metabolism; Low temperature; Metagenome; Microbial Genetics and Genomics; Microbiology; Organic chemicals; Phylogeny; Plasmids; Sequence Analysis, DNA; Soil - chemistry; Soil Microbiology; Soil microorganisms; Studies; Substrate Specificity
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-011-3132-7

Functional screening for lipolytic enzymes at low temperatures resulted in the isolation of the novel cold-active esterases, EstM-N1 and EstM-N2, from a metagenomic DNA library of arctic soil samples. EstM-N1 and EstM-N2 were 395 and 407 amino acids in length, respectively, and showed the highest similarity to class C β-lactamases. However, they shared a relatively low level of sequence similarity (30%) with each other. Phylogenetic analysis of bacterial lipolytic enzymes confirmed that EstM-N1 and EstM-N2 belonged to family VIII of bacterial esterases/lipases. The (His) 6 -tagged esterases were purified to about 99% homogeneity from the soluble fraction of recombinant Escherichia coli cultures. The purified EstM-N1 and EstM-N2 retained more than 50% of maximal activity in the temperature range of 0–35°C, with optimal temperatures of 20°C and 30°C, respectively. Both enzymes preferred the short acyl chains of p -nitrophenyl esters and exhibited very narrow substrate specificity, indicating that they are typical esterases. The β-lactamase activity of EstM-N1 and EstM-N2 was also detected and reached about 31% and 13% of the positive control enzyme, Bacillus cereus β-lactamase, respectively. These first cold-active esterases belonging to family VIII are expected to be useful for potential biotechnological applications as interesting biocatalysts.