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Molecular co-operation between protein PAM and streptokinase for plasmin acquisition by Streptococcus pyogenes
U Ringdahl1, M Svensson, A C Wistedt
1Department of Medical Microbiology, University of Lund, S-223 62 Lund, Sweden.
Abstract:
Bacterial surface-associated plasmin formation is believed to contribute to invasion, although the underlying molecular mechanisms are poorly understood. To define the components necessary for plasmin generation on group A streptococci we used strain AP53 which exposes an M-like protein ("PAM") that contains a plasminogen-binding sequence with two 13-amino acid residues long tandem repeats (a1 and a2). Utilizing an Escherichia coli-streptococcal shuttle vector, we replaced a 29-residue long sequence segment of Arp4, an M-like protein that does not bind plasminogen, with a single (a1) or the combined a1a2 repeats of PAM. When expressed in E. coli, the purified chimeric Arp/PAM proteins both bound plasminogen, as well as plasmin, and when used to transform group A streptococcal strains lacking the plasminogen-binding ability, transformants with the Arp/PAM constructs efficiently bound plasminogen. Moreover, when grown in the presence of plasminogen, both Arp/PAM- and PAM-expressing streptococci acquired surface-bound plasmin. In contrast, plasminogen activation failed to occur on PAM- and Arp/PAM-expressing streptococci carrying an inactivated streptokinase gene: this block was overcome by exogenous streptokinase. Together, these results provide evidence for an unusual co-operation between a surface-bound protein, PAM, and a secreted protein, streptokinase, resulting in bacterial acquisition of a host protease that is likely to spur parasite invasion of host tissues.
Insights
Group A streptococci use surface protein PAM to bind plasminogen, enabling plasmin formation. This process, dependent on streptokinase, facilitates bacterial invasion of host tissues.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Bacterial surface-associated plasmin formation aids invasion but lacks clear molecular understanding.
- Group A streptococci (GAS) utilize surface proteins to interact with host factors.
- Plasminogen binding is a key step in bacterial plasmin generation.
Purpose of the Study:
- To identify the molecular components essential for plasmin generation on GAS.
- To investigate the role of the M-like protein PAM in plasminogen binding and activation.
- To elucidate the interaction between surface proteins and secreted factors in bacterial plasmin acquisition.
Main Methods:
- Genetic engineering of M-like proteins (Arp4 and PAM) in *Escherichia coli* and GAS.
- Construction of chimeric proteins by incorporating plasminogen-binding repeats from PAM into Arp4.
- Assessing plasminogen and plasmin binding to engineered proteins and bacterial strains.
- Evaluating plasminogen activation on streptococcal surfaces with and without functional streptokinase.
Main Results:
- Chimeric Arp/PAM proteins containing PAM repeats bound both plasminogen and plasmin.
- GAS strains expressing Arp/PAM constructs efficiently bound plasminogen.
- Streptococci expressing PAM or Arp/PAM acquired surface-bound plasmin in the presence of plasminogen.
- Plasminogen activation was blocked in strains with inactivated streptokinase but restored by exogenous streptokinase.
Conclusions:
- The M-like protein PAM is crucial for bacterial plasminogen binding.
- Surface-bound PAM and secreted streptokinase cooperate for bacterial plasmin acquisition.
- This mechanism enhances bacterial invasion by facilitating the formation of a host protease on the bacterial surface.