Premium
Purification and characterization of thermophilic and alkalophilic tributyrin esterase from Bacillus strain A30‐1 (ATCC 53841)
Author(s) -
Wang Yongxiang,
Saha Badal C.
Publication year - 1993
Publication title -
journal of the american oil chemists' society
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.512
H-Index - 117
eISSN - 1558-9331
pISSN - 0003-021X
DOI - 10.1007/bf02632155
Subject(s) - tributyrin , ethylenediaminetetraacetic acid , esterase , isoelectric point , chemistry , chromatography , hydrolysis , enzyme assay , enzyme , biochemistry , lipase , organic chemistry , chelation
An extracellular esterase (EC 3.1.1.1) from a thermophilic Bacillus A30‐1 (ATCC 53841) was purified 139‐fold to homogeneity by sodium chloride (6 M) treatment, ammonium sulfate fractionation (30–80%) and phenyl‐Sepharose CL‐6B column chromatography. The native enzyme was a single polypeptide chain with a molecular weight of about 65,000 and an isoelectric point at pH 4.8. The optimum pH for esterase activity was 9.0, and its pH stability range was 5.0–10.5. The optimum temperature for its activity was 60°C. The esterase had a half‐life of 28 h at 50°C, 20 h at 60°C and 16 h at 65°C. It showed the highest activity on tributyrin, with little or no activity toward long‐chain (12–20 carbon) fatty acid esters. The enzyme displayed K m and K cat values of 0.357 mM and 8365/min, respectively, for tributyrin hydrolysis at pH 9.0 and 60°C. Cyclodextrin (α, β, and γ), Ca 2+ , Co 2+ , Mg 2+ and Mn 2+ enhanced the esterase activity, and Zn 2+ and Fe 2+ acted as inhibitors of the enzyme activity. The enzyme activity was not affected by ethylenediaminetetraacetic acid, p‐chloromercuribenzoate and N ‐bromosuccinimide.