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Network of Surface-Displayed Glycolytic Enzymes in Mycoplasma pneumoniae and Their Interactions with Human
Anne Gründel1, Melanie Pfeiffer1, Enno Jacobs1
1TU Dresden, Institute of Medical Microbiology and Hygiene, Dresden, Germany.
Abstract:
In different bacteria, primarily cytosolic and metabolic proteins are characterized as surface localized and interacting with different host factors. These moonlighting proteins include glycolytic enzymes, and it has been hypothesized that they influence the virulence of pathogenic species. The presence of surface-displayed glycolytic enzymes and their interaction with human plasminogen as an important host factor were investigated in the genome-reduced and cell wall-less microorganism Mycoplasma pneumoniae, a common agent of respiratory tract infections of humans. After successful expression of 19 glycolytic enzymes and production of polyclonal antisera, the localization of proteins in the mycoplasma cell was characterized using fractionation of total proteins, colony blot, mild proteolysis and immunofluorescence of M. pneumoniae cells. Eight glycolytic enzymes, pyruvate dehydrogenases A to C (PdhA-C), glyceraldehyde-3-phosphate dehydrogenase (GapA), lactate dehydrogenase (Ldh), phosphoglycerate mutase (Pgm), pyruvate kinase (Pyk), and transketolase (Tkt), were confirmed as surface expressed and all are able to interact with plasminogen. Plasminogen bound to recombinant proteins PdhB, GapA, and Pyk was converted to plasmin in the presence of urokinase plasminogen activator and plasmin-specific substrate d-valyl-leucyl-lysine-p-nitroanilide dihydrochloride. Furthermore, human fibrinogen was degraded by the complex of plasminogen and recombinant protein PdhB or Pgm. In addition, surface-displayed proteins (except PdhC) bind to human lung epithelial cells, and the interaction was reduced significantly by preincubation of cells with antiplasminogen. Our results suggest that plasminogen binding and activation by different surface-localized glycolytic enzymes of M. pneumoniae may play a role in successful and long-term colonization of the human respiratory tract.
Insights
Surface-displayed glycolytic enzymes in Mycoplasma pneumoniae bind human plasminogen, aiding bacterial colonization. This interaction activates plasminogen, facilitating pathogen survival in the respiratory tract.
Area of Science:
- Microbiology
- Molecular Biology
- Pathogenesis
Background:
- Moonlighting proteins, including glycolytic enzymes, can be surface-localized in bacteria and influence virulence.
- Mycoplasma pneumoniae, a respiratory pathogen, lacks a cell wall, making surface protein localization crucial for host interaction.
Purpose of the Study:
- To investigate the surface localization of glycolytic enzymes in Mycoplasma pneumoniae.
- To determine if these surface-expressed enzymes interact with human plasminogen and affect virulence.
Main Methods:
- Expression and antiserum production for 19 glycolytic enzymes.
- Cellular localization studies using fractionation, colony blot, mild proteolysis, and immunofluorescence.
- In vitro assays for plasminogen binding, activation, and fibrinogen degradation.
Main Results:
- Eight glycolytic enzymes (PdhA-C, GapA, Ldh, Pgm, Pyk, Tkt) were confirmed as surface-expressed.
- All eight enzymes bound human plasminogen, with PdhB, GapA, and Pyk activating it to plasmin.
- Surface proteins facilitated binding to lung epithelial cells, reduced by anti-plasminogen antibodies.
Conclusions:
- Surface-localized glycolytic enzymes in M. pneumoniae bind and activate plasminogen.
- This interaction likely contributes to the pathogen's successful colonization of the human respiratory tract.
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