摘要
小白链霉菌(Streptomyces albulus)是ε-聚赖氨酸(ε-poly-L-lysine,ε-PL)的生产菌,且只能在pH 4.0左右环境下大量积累ε-PL,但目前关于小白链霉菌响应低pH胁迫的生理机制研究较少。借助比较转录组学,研究Streptomyces albulus M-Z18响应低pH胁迫的生理机制。在恒化培养的条件下,考察S.albulus M-Z18在pH 5.5和pH 4.0条件下的全局基因转录差异,并对差异基因进行GO和KEGG分析。相比于pH 5.5,pH 4.0条件下共检测出3893个显著差异表达基因,其中有1786个显著上调基因,2107个显著下调基因。KEGG分析发现,S.albulus M-Z18在pH 4.0时增强了糖代谢中糖酵解途径的第三阶段、三羧酸循环、氧化磷酸化途径、戊糖磷酸途径的氧化阶段,减弱了戊糖磷酸途径的非氧化阶段;同时,削弱了细胞壁肽聚糖合成,但增加了肽聚糖交联速度,并增加了细胞膜不饱和脂肪酸、环丙烷脂肪酸的合成,提高了细胞膜脂肪酸的不饱和度;此外,减弱多数氨基酸的合成,增强了碱性氨基酸(组氨酸和赖氨酸)的合成;最后,还增强了蛋白质和DNA修复蛋白的表达。有趣的是,pH 4.0条件下S.albulus M-Z18中所有耐酸系统都不同程度的上调,其中尿素酶系统上调最为显著。低pH环境条件下,S.albulus M-Z18可能是通过增强糖代谢、碱性氨基酸合成、维持细胞壁完整性、增加细胞膜流动性,以及增强蛋白质和DNA修复能力、酸耐受系统来全面应对低pH胁迫。研究结果为进一步探究小白链霉菌响应低pH胁迫的分子机制提供了指导。
ε-Poly-lysine(ε-PL)is a naturally occurring poly(amino acid)consisting of 25-35 L-lysine residues with amide linkages formed between theα-carboxyl group and theε-amino group of the side chain,which possesses excellent antibacterial activity against bacteria and fungi.As a natural preservative,ε-PL is widely used in the food industry.Streptomyces albulus is one of the majorε-PL producers,which can only secret a large amount ofε-PL under environmental conditions around pH 4.0,so it is often subject to acid stress.However,few studies have focused on the physiological mechanisms of S.albulus in responds to low pH stress.Here,we investigated the effect of different pH on the cell growth andε-PL synthesis of S.albulus M-Z18.The results showed that S.albulus M-Z18 could grow normally when pH>3.5,whileε-PL can accumulate when pH≤5,and its production gradually increased with the decreased pH level.Subsequently,the physiological mechanisms of S.albulus M-Z18 in response to low pH stress were investigated by comparative transcriptome analysis.The chemostat culture assay was conducted at different pH(5.5,4.0)by S.albulus M-Z18,and a global transcriptional analysis at pH 5.5 and pH 4.0 were investigated.A total of 3893 significantly differentially expressed genes(DEGs)were detected at pH 4.0 compared to pH 5.5,with 1786 significantly up-regulated DEGs and 2107 down-regulated DEGs,representing 53.97%of the number of genes detected.This result indicated that S.albulus M-Z18 gene expression was significantly altered in response to low pH stress.GO and KEGG analyses of DEGs showed that DEGs were mainly concentrated in biological process(GO analysis)and metabolism(KEGG analysis).Further analysis revealed that genes related to the third stage of the glycolytic pathway,TCA cycle,oxidative phosphorylation pathway and the oxidative stage of the pentose phosphate pathway were significantly upregulated,while genes related to the non-oxidative stage of the pentose phosphate pathway were significantly downregulated.These suggests that the acid tolerance of S.albulus M-Z18 may be enhanced by the improved cell metabolism.The analysis of the cell wall peptidoglycan synthesis pathway showed significant down-regulation of several genes involved in the first phase of cell wall peptidoglycan synthesis,which would lead to an overall limitation of peptidoglycan synthesis.However,some genes involved in peptidoglycan cross-linking and modification were significantly up-regulated at pH 4.0,which would help to increase the rate of cell wall peptidoglycan cross-linking and maintain the integrity of the S.albulus M-Z18 cell wall at low pH.Meanwhile,analysis of the cell membrane fatty acid synthesis pathway showed that genes related to cell membrane unsaturated fatty acid and cyclopropane fatty acid synthesis were significantly up-regulated at pH 4.0,which suggested that the unsaturation of cell membrane fatty acids and the content of cyclopropane fatty acids may be increased.S.albulus M-Z18 may limit the entry of H+into the cell by altering the fatty acid components of the cell membrane,which is conductive to the maintenance of normal physiological functions of S.albulus M-Z18 in a low pH environment.In addition,the analysis of the amino acid synthesis pathway indicated that the synthesis of most amino acids was reduced,but the synthesis of some basic amino acids(histidine and lysine)and substrates of amino acid tolerant systems(glutamate and glutamine)was enhanced,which is advantageous for S.albulus M-Z18 to maintain the stability of intracellular pH.Finally,the changes in genes related to the protection and repair of the intracellular macromolecules under low pH conditions were analyzed.The results showed that molecular chaperone genes,ATP-dependent protease genes,nucleotide excision repair and non-homologous end-joining repair-related genes were significantly up-regulated,indicating that the protein repair capacity and DNA damage repair of S.albulus M-Z18 were enhanced in at low pH,which helped the strain to repair the damage caused by acid stress.The transcriptome data were further retrieved,and S.albulus M-Z18 genome was found to harbor a glutamate decarboxylase system,a glutamate-glutamine transporter protein system,a lysine decarboxylase system,an agmatine deaminase deiminase system,a urease system and an arginine deiminase system.As a result,the glutamate decarboxylase system,glutamate-glutamine transporter protein system,agmatine deiminase system and urease system were all up-regulated to different degrees at pH 4.0.Interestingly,the urease subunitα(UreC)and nickel transporter protein(NixA)were significantly up-regulated by 84.64-fold and 47.97-fold,respectively,which suggests that the urease system may play an important role in the resistance of S.albulus M-Z18 to acid stress.In conclusion,S.albulus M-Z18 may response to low pH stress by enhancing cell metabolism,alkaline amino acid synthesis,cell membrane fluidity,protein and DNA repair capacity,acid tolerance systems as well as maintaining cell wall integrity.The above results provide guidance for further investigation on the molecular mechanism of S.albulus in response to low pH stress.
作者
张越
王开方
潘龙
陈旭升
ZHANG Yue;WANG Kaifang;PAN Long;CHEN Xusheng(School of Biotechnology,Jiangnan University,The Key Laboratory of Industrial Biotechnology,Ministry of Education,Wuxi 214122,China)
出处
《食品与发酵工业》
CAS
CSCD
北大核心
2022年第18期1-10,共10页
Food and Fermentation Industries
基金
江苏省自然科学基金(BK20191332,BK20190585)
国家自然科学基金(31901622,31671846)
国家重点研发计划(2020YFA0907700)。
关键词
酸胁迫
转录组
耐酸机制
小白链霉菌
Ε-聚赖氨酸
acid stress
transcriptome
acid tolerance mechanism
Streptomyces albulus
ε-poly-lysine
