Cloning, purification, and characterization of a cold-adapted esterase produced by Psychrobacter cryohalolentis K5T from Siberian cryopeg
Novototskaya-Vlasova, K; Petrovskaya, L; Yakimov, S; Gilichinsky, D
| HERO ID | 1446593 |
|---|---|
| In Press | No |
| Year | 2012 |
| Title | Cloning, purification, and characterization of a cold-adapted esterase produced by Psychrobacter cryohalolentis K5T from Siberian cryopeg |
| Authors | Novototskaya-Vlasova, K; Petrovskaya, L; Yakimov, S; Gilichinsky, D |
| Journal | FEMS Microbiology Ecology |
| Volume | 82 |
| Issue | 2 |
| Page Numbers | 367-375 |
| Abstract | A psychrotrophic gram-negative bacterium Psychrobacter cryohalolentis K5(T) was previously isolated from a cryopeg within Siberian permafrost and its genome has been completely sequenced. To clone and characterize potential cold-active lipases/esterases produced by P. cryohalolentis K5(T) , we have identified their potential genes by alignment with amino acid sequences of lipases/esterases from related bacteria. One of the targets, EstPc, was cloned and overexpressed in Escherichia coli BL21 (DE3) cells. The recombinant protein was produced with a 6x histidine tag at its C-terminus and purified by nickel affinity chromatography. Purified recombinant protein displayed maximum esterolytic activity with p-nitrophenyl butyrate (C4) as a substrate at 35 °C and pH 8.5. Activity assay conducted at different temperatures revealed that EstPc is a cold-adapted esterase which displayed more than 90% of its maximum activity at 0-5 °C. In contrast to many known cold-active enzymes, it possesses relatively high thermostability, preserving more than 60% of activity after incubation for 1 h at 80 °C. It was activated by Ca(2+) , Mn(2+) , and EDTA whereas Zn(+2) , Cu(+2) , Co(+2) , Ni(+2) , and Mg(+2) inhibited it. Various organic solvents (ethanol, methanol and others) inhibited the enzyme. Most non-ionic detergents, such as Triton X-100 and Tween 20 increased the lipase activity while SDS completely inhibited it. |
| Doi | 10.1111/j.1574-6941.2012.01385.x |
| Pmid | 22486752 |
| Wosid | WOS:000310261900013 |
| Is Certified Translation | No |
| Dupe Override | No |
| Comments | Source: Web of Science WOS:000310261900013 |
| Is Public | Yes |
| Language Text | English |
| Keyword | cold-active esterase; Psychrobacter cryohalolentis; cryopeg; permafrost |