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Immuno-fluorescence Assay of Leptospiral Surface-exposed Proteins
Published on: July 1, 2011
Biological characterization of major polypeptides on the surface of spotted fever group rickettsiae
1Department of Pathology, University of Texas Medical Branch, Galveston 77550.
Abstract:
There are two major polypeptides on the surface of spotted fever group rickettsiae that play important roles as immunogens and possibly relate to the pathogenicity of rickettsial diseases. Digestion with trypsin and staphylococcal V-8 protease shows that the 115-kDa protein of Rickettsia conorii is a trypsin-sensitive, surface-exposed protein. The 135-kDa protein is relatively trypsin resistant. Both proteins resist heat treatment up to 52 degrees C and maintain their antigenic reactivity with monoclonal antibodies when the intact rickettsiae are heated. When rickettsiae are dissolved in SDS-containing sample buffer, these polypeptides lose their antigenic reactivity with monoclonal antibodies after incubation at 45 degrees C for 10 min. Staphylococcal V-8 protease does not appear to affect these polypeptides under the current experimental conditions. Monoclonal antibody to the 115-kDa protein reduces rickettsial attachment. The 115-kDa polypeptide may play a role in rickettsial adhesion to the host cell surface.
Insights
Two major surface proteins of spotted fever group rickettsiae were analyzed. The 115-kDa protein is trypsin-sensitive and may be involved in rickettsial adhesion to host cells.
Area of Science:
- Microbiology
- Immunology
- Molecular Biology
Background:
- Spotted fever group rickettsiae possess surface proteins crucial for immunogenicity and pathogenicity.
- Understanding these proteins is key to addressing rickettsial diseases.
Purpose of the Study:
- To characterize the properties of two major surface polypeptides (115-kDa and 135-kDa) of Rickettsia conorii.
- To investigate their sensitivity to enzymatic digestion and heat, and their role in host cell interaction.
Main Methods:
- Proteolytic digestion using trypsin and staphylococcal V-8 protease.
- Heat treatment of intact and SDS-dissolved rickettsiae.
- Immunological assays using monoclonal antibodies.
- Assessment of rickettsial attachment to host cells.
Main Results:
- The 115-kDa protein is trypsin-sensitive and surface-exposed, while the 135-kDa protein is relatively trypsin-resistant.
- Both proteins retain antigenic reactivity after heat treatment of intact rickettsiae up to 52°C.
- Antigenic reactivity is lost upon heating SDS-dissolved polypeptides at 45°C.
- Staphylococcal V-8 protease had no significant effect under tested conditions.
- A monoclonal antibody against the 115-kDa protein inhibited rickettsial attachment.
Conclusions:
- The 115-kDa polypeptide is a surface-exposed, trypsin-sensitive protein potentially involved in Rickettsia conorii adhesion.
- These surface proteins are important targets for understanding rickettsial pathogenicity and host interactions.
