研究报告

黑曲霉脂肪酶tglE的基因克隆与生化特征解析

  • 丛珊滋 ,
  • 田康明 ,
  • 张新 ,
  • 路福平 ,
  • 王正祥
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  • 1 (天津科技大学 生物工程学院,工业发酵微生物教育部重点实验室,天津,300457)
    2 (天津科技大学 化工与材料学院,天津,300457)
    3 (北京农学院,农产品有害微生物及农残安全检测与控制北京市重点实验室,北京,102206)
博士研究生(路福平教授为通讯作者,E-mail:lfp@tust.edu.cn)。

收稿日期: 2019-03-19

  网络出版日期: 2019-11-15

基金资助

政府间国际合作重点项目(2018YFE0100400);天津市杰出津门学者计划

Gene cloning and biochemical characterization of Aspergillus niger lipase tglE

  • CONG Shanzi ,
  • TIAN Kangming ,
  • ZHANG Xin ,
  • LU Fuping
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  • 1 (Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin Universityof Science and Technology, Tianjin 300457,China)
    2 (College of Chemical Engineering and Materials Science, Tianjin University of Science and Technology, Tianjin 300457,China)
    3 (Beijing Key Laboratory of Detection and Control of Spoilage Microorganisms and Pesticide Residues in AgriculturalProducts, Beijing University of Agriculture, Beijing 102206, China)

Received date: 2019-03-19

  Online published: 2019-11-15

摘要

从黑曲霉基因组中克隆了一种新的脂肪酶基因tglE,并在毕赤酵母中成功表达。重组酶具有典型的脂肪酶活性,其最适pH为7.0,最适温度为40 ℃;在30~60 ℃或pH 6.5~9.0该酶的酶活力保持在60%以上;在pH 6.0~8.0或20~30 ℃均较为稳定。Cu2+、Mn2+、Ca2+、K+、Mg2+和Sn2+tglE有明显的激活作用;Fe3+、Zn2+和EDTA对重组酶的酶活抑制作用明显,特别是EDTA可抑制重组酶90%的活。tglE对橄榄油和C4底物的作用最强,对棕果油和C16底物的作用最弱。以C4为底物,tglEKm值为14.40 mmol/L,Vmax为46.72 mmol/(mL·h)。tglE具有催化辛酸和乙醇生成辛酸乙酯的酯化活性。

本文引用格式

丛珊滋 , 田康明 , 张新 , 路福平 , 王正祥 . 黑曲霉脂肪酶tglE的基因克隆与生化特征解析[J]. 食品与发酵工业, 2019 , 45(21) : 1 -7 . DOI: 10.13995/j.cnki.11-1802/ts.020595

Abstract

The novel lipase gene (tglE) from Aspergillus niger was cloned and expressed in Pichia pastoris. The recombinant TglE had typical lipase activity with the maximal activity at 40 ℃ and pH 7.0. The enzyme retained over 60% activities at 30 ℃-60 ℃ or pH 6.5-9.0 and was stable at 20-30 ℃ or pH 6.0-8.0. The catalytic activity of TglE was significantly improved by Cu2+, Mn2+, Ca2+, K+, Mg2+, and Sn2+ but inhibited by Fe3+, Zn2+ and EDTA. The prefered substrates for tglE were chemically synthesized pNPB and natural olive oil. The Km and Vmax for pNPB were 14.40 mmol/L and 46.72 mmol/(mL·h), respectively. In addition to the hydrolysis activity, tglE exhibited significant esterification efficiency for ethyl caprylate synthesis from octanoic acid and ethanol.

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