Factor H Is Bound by Outer Membrane-Displayed Carbohydrate Metabolism Enzymes of Extraintestinal Pathogenic Escherichia coli and Contributes to Opsonophagocytosis Resistance in Bacteria

Frontiers in Cellular and Infection Microbiology

Qianwen Gong¹  Xuhang Wang¹  Rui Chen¹  Feng Xue²  Fang Tang²  Yu Sun²  Xiangkai Zhuge³  Bin Xu⁴  Jianjun Dai⁵ 

[1] Key Laboratory of Animal Bacteriology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, China;MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China;Key Laboratory of Animal Bacteriology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, China;MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China;Key Laboratory of Animal Bacteriology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, China;Department of Nutrition and Food Hygiene, School of Public Health, Nantong University, Nantong, China;MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China;Key Laboratory of Animal Bacteriology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, China;National Research Center of Veterinary Biologicals Engineering and Technology, Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Nanjing, China;MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China;Key Laboratory of Animal Bacteriology, Ministry of Agriculture, Nanjing Agricultural University, Nanjing, China;School of Life Science and Technology, China Pharmaceutical University, Nanjing, China;

关键词: complement system;    factor H;    Escherichia coli;    carbohydrate metabolism enzymes;    opsonophagocytosis;   

DOI  :  10.3389/fcimb.2020.592906

来源: Frontiers

【 摘 要 】

Extraintestinal pathogenic Escherichia coli (ExPEC) causes bloodstream infections in humans and animals. Complement escape is a prerequisite for bacteria to survive in the bloodstream. Factor H (FH) is an important regulatory protein of the complement system. In this study, ExPEC was found to bind FH from serum. However, the mechanisms of ExPEC binding to FH and then resistance to complement-mediated attacks remain unclear. Here, a method that combined desthiobiotin pull-down and liquid chromatography-tandem mass spectrometry was used to identify the FH-binding membrane proteins of ExPEC. Seven identified proteins, which all were carbohydrate metabolic enzymes (CMEs), including acetate kinase, fructose-bisphosphate aldolase, fumarate reductase flavoprotein subunit, L-lactate dehydrogenase, dihydrolipoamide dehydrogenase, phosphoenolpyruvate synthase, and pyruvate dehydrogenase, were verified to recruit FH from serum using GST pull-down and ELISA plate binding assay. The ELISA plate binding assay determined that these seven proteins bind to FH in a dose-dependent manner. Magnetic beads coupled with any one of seven proteins significantly reduced the FH recruitment of ExPEC (p < 0.05) Subsequently, immunofluorescence, colony blotting, and Western blotting targeting outer membrane proteins determined that these seven CMEs were located on the outer membrane of ExPEC. Furthermore, the FH recruitment levels and C3b deposition levels on bacteria were significantly increased and decreased in an FH-concentration-dependent manner, respectively (p < 0.05). The FH recruitment significantly enhanced the ability of ExPEC to resist the opsonophagocytosis of human macrophage THP-1 in an FH-concentration-dependent manner (p < 0.05), which revealed a new mechanism for ExPEC to escape complement-mediated killing. The identification of novel outer membrane-displayed CMEs which played a role in the FH recruitment contributes to the elucidation of the molecular mechanism of ExPEC pathogenicity.

【 授权许可】

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