食品与发酵工业 >

2023 , Vol. 49 >Issue 13: 40 - 48

DOI: https://doi.org/10.13995/j.cnki.11-1802/ts.033466

研究报告

一种海洋几丁质酶产生菌的筛选及酶学性质研究

  • 严佳佳 ,
  • 尤田 ,
  • 张雪梅 ,
  • 李欣格 ,
  • 薛鲜丽 ,
  • 王德培
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  • 1(天津科技大学 生物工程学院,天津,300457)
    2(教育部工业微生物重点实验室,天津,300457)
    3(天津市微生物代谢与发酵过程控制技术工程中心,天津,300457)
第一作者:硕士研究生(王德培教授为通信作者,E-mail:wangdp@tust.edu.cn)

收稿日期: 2022-08-29

  修回日期: 2022-09-23

  网络出版日期: 2023-08-07

基金资助

天津市自然科学基金重点项目(20JCZDJC00140);国家自然科学基金青年科学基金项目(31902193)

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Screening of a marine chitinase-producing bacterium and its enzymatic properties

  • YAN Jiajia ,
  • YOU Tian ,
  • ZHANG Xuemei ,
  • LI Xinge ,
  • XUE Xianli ,
  • WANG Depei
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  • 1(College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China)
    2(Laboratory of Industrial Fermentation Microbiology of Ministry of Education, Tianjin 300457, China)
    3(Tianjin Microbial Metabolism and Fermentation Process Control Technology Engineering Center, Tianjin 300457, China)

Received date: 2022-08-29

  Revised date: 2022-09-23

  Online published: 2023-08-07

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摘要

海洋是自然界几丁质主要来源地,滋养出种类繁多可降解几丁质的微生物,因此从海洋中筛选产高活性几丁质酶的细菌,是获得高产几丁质酶微生物的有效途径。该文以胶体几丁质为唯一碳源,从渤海滩涂筛选到一株具有降解几丁质特性的细菌Y-8,对其进行鉴定并研究其几丁质酶酶学性质。Y-8菌株经分子鉴定为副溶血性弧菌(Vibrio parahaemolyticus),发酵上清液经SDS-PAGE及蛋白质谱分析,发现其存在2种几丁质酶,分别为Chi1几丁质外切酶,氨基酸残基数为848,理论分子质量为87.6 kDa;Chi2几丁质内切酶,氨基酸残基数为1 054,理论分子质量为112.9 kDa。Y-8所产生的几丁质酶能够高效降解胶体几丁质,获得单一产物N-乙酰氨基葡萄糖,其最适反应温度为55 ℃,45 ℃保温1 h,仍能保持70%以上活性;最适pH为6.0,在pH 4.0~9.0、37 ℃保温1 h后相对酶活性保留55%以上,具有较好的稳定性。10 mmol/L的Mn2+能使几丁质酶活性提高362%,而EDTA以及SDS、吐温-20、吐温-80对酶活力具有抑制作用。

关键词： 菌株筛选; 副溶血性弧菌; 几丁质; 几丁质酶; 酶学性质

本文引用格式

严佳佳 , 尤田 , 张雪梅 , 李欣格 , 薛鲜丽 , 王德培 . 一种海洋几丁质酶产生菌的筛选及酶学性质研究[J]. 食品与发酵工业, 2023 , 49(13) : 40 -48 . DOI: 10.13995/j.cnki.11-1802/ts.033466

Abstract

Ocean is a major source of chitin in nature, which encourages a wide variety of microorganisms capable of degrading chitinase. Therefore, screening bacteria with high activity of chitinase from the ocean is an effective way to obtain microorganisms with high yield of chitinase. In this paper, a bacterial strain Y-8, which can degrade chitin, was screened from the sea crab and its habitant-Bohai mudflats, with colloidal chitin as the sole carbon source, and its identification and the enzymatic properties of its chitinase were studied. Strain Y-8 was identified as Vibrio parahaemolyticus by molecular identification. SDS-PAGE and protein spectrum analysis of the fermentation broth supernatant revealed the presence of two chitinases. Chi1 chitinase, with amino acid residue of 848, has theoretical molecular weight of 87.6 kDa. Chi2 chitin endonuclease has a residual amino acid number of 1 054 and a theoretical molecular weight of 112.9 kDa. The chitinase produced by Y-8 could effectively degrade colloidal chitin to obtain a single product N-acetyl glucosamine. The optimal reaction temperature was 55 ℃, and the enzyme retained more than 70% activity after 1 h at 45 ℃. The optimum pH was 6.0, and the enzyme retained more than 55% relative enzyme retained after 1 h at pH 4.0-9.0 37 ℃, showing good stability. 10 mmol/L Mn2+ could increase chitinase activity by 362%, while EDTA, SDS, tween-20, tween-80 could inhibit chitinase activity.

Key words： strain screening; Vibrio parahaemolyticus; chitin; chitinase; enzymatic properties

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