Comparative Proteome Analysis of Shewanella putrefaciens WS13 Mature Biofilm Under Cold Stress
Comparative Proteome Analysis of Shewanella putrefaciens WS13 Mature Biofilm Under Cold Stress
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DOI：
10.3389/fmicb.2020.01225
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发表时间：
2020-06
影响因子：
5.2
通讯作者：
Jun-li Yan;Jing Xie
中科院分区：
文献类型：
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作者：
Jun-li Yan;Jing Xie
腐败希瓦氏菌（Shewanellaputrefaciens）是世界范围内冷藏期间主要的海产品腐败微生物。这种细菌可以附着在生物/非生物表面，形成生物膜，导致海产品质量下降和保质期缩短。对S.腐败菌生物膜的形成尚未描述。结晶紫染色结合共聚焦激光扫描显微镜（CLSM）用于研究导致在4、15和30°C下建立成熟生物膜的事件的顺序。此外，成熟的生物膜的主要化学成分确定的拉曼光谱（RM），而比较蛋白质组学分析被用来量化代谢途径的变化，并找出潜在的蛋白质决定因素。成熟生物膜的物理尺寸，即，生物量、生物体积和平均厚度在4°C下高于15和30°C。通过RM测量的蛋白质的变化证实了蛋白质在成熟生物膜形成过程中的重要性。比较蛋白质组学分析表明，铁载体和铁螯合物转运蛋白在成熟的生物膜形成过程中下调。上述下调的蛋白质参与促进铁储存，以响应对代谢能量的更高需求，而硫中继系统、嘧啶代谢和嘌呤代谢的上调蛋白质与细菌适应性有关。与冷胁迫相关的蛋白质合成增加，参与氨酰-tRNA生物合成的蛋白质表达上调，而参与氨肽酶活性的蛋白质表达下调。蛋白质水解的能量减少，参与焦磷酸酶活性的蛋白质上调。此外，还发现了胞外eDNA，它可能在维持成熟S.低温胁迫下腐败菌生物膜结构的变化。这项工作提供了一个更好的理解蛋白质在成熟生物膜中的作用。此外，还探讨了一种耐冷腐败菌在低温下的生物膜形成机理，为海产品保鲜和加工过程中生物膜的控制提供了理论依据。
Worldwide, Shewanella putrefaciens is the predominant seafood spoilage microorganism during cold storage. This bacterium can attach to biotic/abiotic surfaces to form biofilms which contribute to seafood quality degradation and shelf-life reduction. The mechanism of S. putrefaciens biofilm formation is not yet described. Crystal violet staining in combination with confocal laser scanning microscopy (CLSM) was used to study the sequence of events leading to the establishment of a mature biofilm at 4, 15, and 30°C. In addition, the main chemical constituents of the mature biofilm were determined by Raman spectroscopy (RM), whereas, comparative proteomic analysis was used to quantify changes in metabolic pathways and to find out underlying protein determinants. The physical dimensions of the mature biofilm, i.e., biomass, biovolume, and mean thickness, were higher at 4°C when compared to 15 and 30°C. The variations of proteins measured by RM confirmed the importance of proteins during the formation of a mature biofilm. Comparative proteomic analysis showed that siderophore and iron chelate transport proteins were down-regulated during mature biofilm formation. The down-regulated aforementioned proteins are involved in promoting iron storage in response to a higher demand for metabolic energy, whereas, the upregulated proteins of the sulfur relay system, pyrimidine metabolism, and purine metabolism are related to bacterial adaptability. Synthesis of proteins related to cold stress was increased and proteins involved in aminoacyl-tRNA biosynthesis were up-regulated, whereas, proteins involved in aminopeptidase activity were down-regulated. Proteolysis to scavenge energy was reduced as proteins involved in pyrophosphatase activity were up-regulated. Also extracellular eDNA was found which may play an important role in maintaining the stability of mature S. putrefaciens biofilm structures under cold stress. This work provides a better understanding of the role of proteins in mature biofilms. In addition, the biofilm formation mechanism of a psychrotrophic spoilage bacterial species at low temperature is explored, which may contribute to generating biofilm controlling strategies during seafood preservation and processing.