Biological Science Program and Center of Excellence for Innovation in Chemistry (PERCH-CIC), Faculty of Science, Burapha University, Bangsaen, Chonburi 20131, Thailand.
Appl Biochem Biotechnol. 2010 Nov;162(5):1362-76. doi: 10.1007/s12010-010-8928-x. Epub 2010 Feb 24.
The benzene tolerant Acinetobacter baylyi isolated from marine sludge in Angsila, Thailand could constitutively secrete lipolytic enzymes. The enzyme was successfully purified 21.89-fold to homogeneity by ammonium sulfate precipitation and gel-permeable column chromatography with a relative molecular mass as 30 kDa. The enzyme expressed maximum activity at 60 degrees C and pH 8.0 with p-nitrophenyl palmitate as a substrate and found to be stable in pH and temperature ranging from 6.0-9.0 to 60-80 degrees C, respectively. A study on solvent stability revealed that the enzyme was highly resisted to many organic solvents especially benzene and isoamyl alcohol, but 40% inhibited by decane, hexane, acetonitrile, and short-chain alcohols. Lipase activity was completely inhibited in the presence of Fe(2+), Mn(2+), EDTA, SDS, and Triton X-100 while it was suffered detrimentally by Tween 80. The activity was enhanced by phenylmethylsulfonyl fluoride (PMSF), Na(+), and Mg(2+) and no significant effect was found in the presence of Ca(2+) and Li(+). Half of an activity was retained by Ba(2+), Ag(+), Hg(+), Ni(2+), Zn(2+), and DTT. The enzyme could hydrolyze a wide range of p-nitrophenyl esters, but preferentially medium length acyl chains (C(8)-C(12)). Among natural oils and fats, the enzyme 11-folds favorably catalyzed the hydrolysis of rice bran oil, corn oil, sesame oil, and coconut oil in comparison to palm oil. Moreover, the transesterification activity of palm oil to fatty acid methyl esters (FAMEs) revealed 31.64 +/- 1.58% after 48 h. The characteristics of novel A. baylyi lipase, as high temperature stability, organic solvent tolerance, and transesterification capacity from palm oil to FAMEs, indicate that it could be a vigorous biocatalyzer in the prospective fields as bioenergy industry or even in organic synthesis and pharmaceutical industry.
从泰国安锡拉海洋淤泥中分离到的耐苯的不动杆菌（Acinetobacter baylyi）可以组成性分泌脂肪酶。该酶通过硫酸铵沉淀和凝胶渗透柱层析成功地纯化了 21.89 倍，达到均一性，相对分子质量为 30 kDa。该酶在以对硝基苯棕榈酸酯为底物时，在 60°C 和 pH8.0 下表达最大活性，并且在 pH6.0-9.0 和温度 60-80°C 范围内分别稳定。溶剂稳定性研究表明，该酶对许多有机溶剂具有高度抗性，特别是苯和异戊醇，但被癸烷、己烷、乙腈和短链醇抑制 40%。脂肪酶活性在存在 Fe(2+)、Mn(2+)、EDTA、SDS 和 Triton X-100 时完全被抑制，而在 Tween 80 存在时则受到损害。苯甲基磺酰氟（PMSF）、Na(+)和 Mg(2+) 可增强其活性，而 Ca(2+)和 Li(+) 的存在则没有明显影响。一半的活性保留在 Ba(2+)、Ag(+)、Hg(+)、Ni(2+)、Zn(2+)和 DTT 中。该酶可以水解广泛的对硝基苯酯，但优先水解中长链酰基（C(8)-C(12)）。在天然油脂中，该酶对米糠油、玉米油、芝麻油和椰子油的水解作用比棕榈油有利 11 倍。此外，棕榈油到脂肪酸甲酯（FAMEs）的转酯化活性在 48 小时后显示出 31.64 +/- 1.58%。新型不动杆菌脂肪酶的特性，如高温稳定性、有机溶剂耐受性和从棕榈油到 FAMEs 的转酯化能力，表明它可能成为生物能源工业甚至有机合成和制药工业等有前途领域的有力生物催化剂。
