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Biochemical and Structural Characterization of the Carbohydrate Transport Substrate-binding-protein SP0092
Published on: October 2, 2017
Identification and characterization of human surfactant protein A binding protein of Mycoplasma pneumoniae
T R Kannan1, D Provenzano, J R Wright
1Department of Microbiology and Immunology, The University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, Texas 78229-3900, USA.
Abstract:
Mycoplasma pneumoniae infections represent a major primary cause of human respiratory diseases, exacerbate other respiratory disorders, and are associated with extrapulmonary pathologies. Cytadherence is a critical step in mycoplasma colonization, aided by a network of mycoplasma adhesins and cytadherence accessory proteins which mediate binding to host cell receptors. Furthermore, the respiratory mucosa is enriched with extracellular matrix components, including surfactant proteins, fibronectin, and mucin, which provide additional in vivo targets for mycoplasma parasitism. In this study we describe interactions between M. pneumoniae and human surfactant protein-A (hSP-A). Initially, we found that viable M. pneumoniae cells bound to immobilized hSP-A in a dose- and calcium (Ca(2+))-dependent manner. Mild trypsin treatment of intact mycoplasmas reduced binding markedly (80 to 90%) implicating a surface-associated mycoplasma protein(s). Using hSP-A-coupled Sepharose affinity chromatography and polyacrylamide gel electrophoresis, we identified a 65-kDa hSP-A binding protein of M. pneumoniae. The presence of Ca(2+) enhanced binding of the 65-kDa protein to hSP-A, which was reduced by the divalent cation-chelating agent, EDTA. The 65-kDa hSP-A binding protein of M. pneumoniae was identified by sequence analysis as a novel protein (MPN372) possessing a putative S1-like subunit of pertussis toxin at the amino terminus (amino acids 1 to 226), with the remaining amino acids (227 to 591) exhibiting no homology with other subunits of pertussis toxin, other known toxins, or any reported proteins. Recombinant MPN372 (MPN372) bound to hSP-A in a dose-dependent manner, which was markedly reduced by preincubation with mouse recombinant MPN372 antisera. Also, adherence of viable M. pneumoniae cells to hSP-A was inhibited by recombinant MPN372 antisera, demonstrating that MPN372, a previously designated hypothetical protein, is surface exposed and mediates mycoplasma attachment to hSP-A.
Insights
Mycoplasma pneumoniae uses a novel 65-kDa protein, MPN372, to attach to human surfactant protein-A (hSP-A). This interaction is calcium-dependent and crucial for colonization of the respiratory tract.
Area of Science:
- Microbiology
- Infectious Diseases
- Molecular Biology
Background:
- Mycoplasma pneumoniae causes respiratory diseases and attaches to host cells via adhesins.
- Respiratory mucosa contains extracellular matrix proteins like surfactant protein-A (hSP-A) that can be targeted by mycoplasma.
- Understanding M. pneumoniae adherence mechanisms is key to combating respiratory infections.
Purpose of the Study:
- To investigate the interaction between Mycoplasma pneumoniae and human surfactant protein-A (hSP-A).
- To identify and characterize the M. pneumoniae protein responsible for binding to hSP-A.
Main Methods:
- Binding assays using viable M. pneumoniae and immobilized hSP-A.
- Affinity chromatography coupled with gel electrophoresis to identify binding proteins.
- Sequence analysis to identify the novel protein (MPN372).
- Recombinant protein expression and binding studies, including inhibition assays.
Main Results:
- Viable M. pneumoniae bound to hSP-A in a dose- and calcium-dependent manner.
- A 65-kDa protein (MPN372) was identified as the primary hSP-A binding protein.
- MPN372 possesses a novel structure with a putative S1-like subunit and mediates M. pneumoniae attachment to hSP-A.
- Antibodies against MPN372 inhibited M. pneumoniae adherence to hSP-A.
Conclusions:
- MPN372 is a surface-exposed protein mediating Mycoplasma pneumoniae attachment to human surfactant protein-A.
- This interaction is a significant factor in the colonization of the respiratory tract by M. pneumoniae.
- MPN372 represents a potential target for therapeutic interventions against M. pneumoniae infections.
