1. [Cloning, heterologous expression and characterization of a thermostable esterase from Bacillus sp. HJ14 for diethyl-phthalate degradation].
- Author
-
Peng Z, Ding J, Yang Y, Li J, Mu Y, and Huang Z
- Subjects
- Amino Acid Sequence, Bacillus chemistry, Bacillus genetics, Bacterial Proteins genetics, Biodegradation, Environmental, Butyrates chemistry, Butyrates metabolism, Enzyme Stability, Esterases genetics, Hydrogen-Ion Concentration, Mass Spectrometry, Phthalic Acids chemistry, Phylogeny, Sequence Homology, Amino Acid, Substrate Specificity, Temperature, Bacillus enzymology, Bacterial Proteins chemistry, Bacterial Proteins metabolism, Cloning, Molecular, Esterases chemistry, Esterases metabolism, Phthalic Acids metabolism
- Abstract
Objective: A thermostable esterase EstZ1 from Bacillus sp. HJ14 able to degrade diethyl-phthalate (DEP) was heterologously expressed in Escherichia coli BL21(DE3) and characterized., Methods: Full-length EstZ1 was obtained based on specific amplification and genome sequencing, and amino acid sequence of EstZ1 was analyzed. EstZ1 was expressed in Escherichia coli BL21(DE3) using the pEASY-E2 expression system. EstZ1 was purified to electrophoretic homogeneity by Ni2+-NTA metal chelating affinity chromatography, and the enzyme was characterized. The degradation products from DEP were detected by high-pressure liquid chromatography and electrospray ionization mass spectrometry., Results: The 903 bp full-length EstZ1 encoded 300 amino acid residues (EstZ1:33.84 kDa). EstZ1 showed the highest identity of 98% with hormone-sensitive lipase (HSL)-like family in NCBI databases. The optimal temperature and pH was 50℃ and 9.0, respectively, with p-NP butyrate as the best substrate. Meanwhile, it was stable between 40 and 70℃, pH 7.0 to 9.5. Most of metal ions, chemical agents had little impact. DEP could partially be degraded by EstZ1 to its corresponding monoalkyl and alcohol., Conclusion: Our findings may serve as reference for phthalate esters degradation.
- Published
- 2016