基于两种预处理方法的白酒酒醅微生物宏蛋白质组学比较

Comparative Metaproteomics of Microbial Communities in Fermented Grains for Baijiu Production Using Two Pretreatment Methods

分别采用离心-超声波破碎法(centrifugation-ultrasonic disruption,C-UD)和液氮研磨-超声波破碎法(liquid nitrogen grinding-ultrasonic disruption,LNG-UD)预处理浓香型白酒酒醅样品,采用Tris/酚提取-甲醇/醋酸铵沉淀法提取微生物蛋白质,利用非标记定量蛋白质组学技术比较两种预处理法获得的酒醅微生物群落结构差异。结果表明:两种预处理法提取的酒醅蛋白质含量差异不显著,但C-UD处理十二烷基硫酸钠-聚丙烯酰胺凝胶电泳的蛋白条带数量更丰富且清晰。基于C-UD法获得的酒醅肽段及蛋白质数量高于LNG-UD法,且蛋白质鉴定结果的评分较高。两种预处理法获得的酒醅微生物种类及其群落结构有一定差异,且两者均与同一酒醅样品的基因组扩增子测序结果存在差异,提示整合多组学研究的必要性。研究结果有助于进一步优化酒醅微生物蛋白质的提取方法,为酿酒微生物宏蛋白质组学的深入研究奠定一定基础。

In this study, microbial proteins were extracted by Tris/phenol extraction-methanol/ammonium acetate precipitation from fermented grains for nongxiangxing baijiu production pretreated by centrifugation-ultrasonic disruption(C-UD) or liquid nitrogen grinding-ultrasonic disruption(LNG-UD) and the structure of the microbial communities obtained by the two pretreatment methods was evaluated by using label-free quantitative proteomics. The results showed that there was no significant difference between the protein contents extracted by both pretreatment methods. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) showed that the C-UD extract contained more and clearer protein bands. The number of identified peptides and proteins obtained by C-UD was higher than that obtained by LNG-UD, and the Score Sequest HT of protein identification was higher. The microbial species and community structure obtained by the two pretreatment methods were significantly different, and both were significantly different from the results obtained by genomic amplicon sequencing for the same samples, implying the importance of multi-omic designs for studying the microbial community in fermented grains. The results of the present study will help to optimize the extraction of microbial proteins from fermented grains for baijiu production, and lay a foundation for further understanding baijiu fermentation and improving baijing quality.