金黄杆菌ZHDP1菌株基因组分析及其蛋白酶活性特征与产酶优化

Genome analysis, protease characterization, and enzyme production optimization of Chryseobacterium sp. ZHDP1

摘要:

【背景】蛋白酶活性研究对于理解金黄杆菌属(Chryseobacterium)菌株的生态角色、工业价值及潜在致病机理十分重要。【目的】分析一株细菌的新型基因组序列，以此推断其中潜在的蛋白酶基因，并对其蛋白酶活性进行验证与优化，为后续研究提供数据支撑。【方法】利用高通量测序技术对菌株基因组序列进行测定与组装，并利用单因素优化方法对菌株的蛋白酶作用条件与产酶条件进行优化。【结果】分离得到一株蛋白酶活性菌株，通过分子生物学手段鉴定为金黄杆菌属菌株，并命名为Chryseobacterium sp. ZHDP1。该菌株基因组大小为4 917 748 bp，GC含量为35.95%，平均核苷酸相似性和DNA-DNA杂交指数分别为91.39和47.8。该菌株基因组中发现超过20条蛋白酶基因，上清液中也检测到蛋白酶活性，其最适反应温度和pH值分别为50 ℃和7.0。Zn²⁺、Mn²⁺和Cr₂O₇^2-对蛋白酶活性有强烈抑制作用。ZHDP1菌株在培养温度35 ℃、培养时间35 h、接种量4%、碳源和氮源为玉米粉及摇床转速100 r/min的条件下产酶量最高。【结论】本研究提供了ZHDP1菌株的完整基因组序列，并确定了该菌株的蛋白酶相关特性，为该菌株生态功能、致病机制及应用价值的深入研究奠定了基础。

Abstract:

[Background] Characterizing the protease activity is essential for revealing the potential ecological roles, industrial values, and pathogenic mechanism of Chryseobacterium. [Objective] This paper aims to analyze the genome sequence, characterize the protease, and optimize the enzyme production conditions of a novel Chryseobacterium strain, thereby providing the data support for subsequent research. [Methods] The genome sequence of the strain was obtained by high-throughput sequencing technology, and the conditions of protease activity and enzyme production were optimized by single factor tests. [Results] A strain, Chryseobacterium sp. ZHDP1, was isolated in this work. The genome of this strain had a length of 4 917 748 bp and the GC content of 35.95%. The average nucleotide identity (ANI) and DNA-DNA hybridization (DDH) indexes of the genome were 91.39 and 47.8, respectively. More than 20 protease genes were identified in the genome, and protease activity was detected in the supernatant of the fermentation broth. The protease had the optimum performance at 50 ℃ and pH 7.0 and was strongly inhibited by Zn²⁺, Mn²⁺, and Cr₂O₇^2-. The optimal conditions for the protease production of Chryseobacterium sp. ZHDP1 were as follows: culture temperature of 35 ℃, culture time of 35 h, inoculum amount of 4%, carbon source and nitrogen source of corn flour, and shaking speed of 100 r/min. [Conclusion] This study reveals the full-length genome sequence and the protease characteristics of Chryseobacterium sp. ZHDP1, which lays a foundation for the in-depth study of this strain.