Gründel Anne, Friedrich Kathleen, Pfeiffer Melanie, Jacobs Enno, Dumke Roger
TU Dresden, Institute of Medical Microbiology and Hygiene, Dresden, Germany.
PLoS One. 2015 May 15;10(5):e0126600. doi: 10.1371/journal.pone.0126600. eCollection 2015.
The dual role of glycolytic enzymes in cytosol-located metabolic processes and in cell surface-mediated functions with an influence on virulence is described for various micro-organisms. Cell wall-less bacteria of the class Mollicutes including the common human pathogen Mycoplasma pneumoniae possess a reduced genome limiting the repertoire of virulence factors and metabolic pathways. After the initial contact of bacteria with cells of the respiratory epithelium via a specialized complex of adhesins and release of cell-damaging factors, surface-displayed glycolytic enzymes may facilitate the further interaction between host and microbe. In this study, we described detection of the four subunits of pyruvate dehydrogenase complex (PDHA-D) among the cytosolic and membrane-associated proteins of M. pneumoniae. Subunits of PDH were cloned, expressed and purified to produce specific polyclonal guinea pig antisera. Using colony blotting, fractionation of total proteins and immunofluorescence experiments, the surface localization of PDHA-C was demonstrated. All recombinant PDH subunits are able to bind to HeLa cells and human plasminogen. These interactions can be specifically blocked by the corresponding polyclonal antisera. In addition, an influence of ionic interactions on PDHC-binding to plasminogen as well as of lysine residues on the association of PDHA-D with plasminogen was confirmed. The PDHB subunit was shown to activate plasminogen and the PDHB-plasminogen complex induces degradation of human fibrinogen. Hence, our data indicate that the surface-associated PDH subunits might play a role in the pathogenesis of M. pneumoniae infections by interaction with human plasminogen.
本文描述了糖酵解酶在位于胞质溶胶的代谢过程以及对毒力有影响的细胞表面介导功能中的双重作用，涉及多种微生物。包括常见人类病原体肺炎支原体在内的柔膜菌纲无细胞壁细菌，其基因组减少，限制了毒力因子和代谢途径的种类。细菌通过黏附素的特殊复合物与呼吸道上皮细胞初次接触并释放细胞损伤因子后，表面展示的糖酵解酶可能促进宿主与微生物之间的进一步相互作用。在本研究中，我们描述了在肺炎支原体的胞质和膜相关蛋白中检测到丙酮酸脱氢酶复合物（PDHA - D）的四个亚基。克隆、表达并纯化了PDH的亚基，以产生特异性的豚鼠多克隆抗血清。通过菌落印迹、总蛋白分级分离和免疫荧光实验，证实了PDHA - C的表面定位。所有重组PDH亚基都能够与HeLa细胞和人纤溶酶原结合。这些相互作用可被相应的多克隆抗血清特异性阻断。此外，证实了离子相互作用对PDHC与纤溶酶原结合的影响以及赖氨酸残基对PDHA - D与纤溶酶原结合的影响。已证明PDHB亚基可激活纤溶酶原，且PDHB - 纤溶酶原复合物可诱导人纤维蛋白原降解。因此，我们的数据表明，表面相关的PDH亚基可能通过与人纤溶酶原相互作用在肺炎支原体感染的发病机制中发挥作用。
