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Phosphorylase-cross-reactive antibodies evoked by streptococcal M protein
J B Dale1, H S Courtney, M Kotb
1Veterans Administration Medical Center, Memphis, Tennessee.
Abstract:
Rabbit antisera evoked by type 5 streptococcal M protein (M5) were screened by enzyme-linked immunosorbent assay (ELISA) for immunological cross-reactivity with purified rabbit muscle phosphorylases a and b. Of 10 pep M5 antisera tested, 3 showed significant cross-reactivity with both forms of the enzyme. ELISA inhibition studies using one of the pep M5 antisera showed that all of the phosphorylase b antibodies were inhibited by pep M5, the immunogen, and phosphorylase b, the ELISA antigen. All of the antibodies were also inhibited by pep M6 and pep M19, but not by pep M24, indicating that the cross-reactive epitopes were shared by multiple serotypes of M protein. Western blot (immunoblot) analyses showed that pep M5 antisera reacted strongly with the subunit of phosphorylase b. In addition, purified phosphorylase partially inhibited the binding of pep M5 antibodies to a 95-kilodalton protein of human myocardium. One of the three cross-reactive pep M5 antisera inhibited the enzymatic activity of phosphorylase a in a dose-related fashion, reaching a maximum inhibition of 75%. The enzymatic activity in the presence of antibody was totally restored when the antiserum was first incubated with pep M5.
Insights
Antibodies against streptococcal M protein (M5) cross-reacted with muscle phosphorylase enzymes. This cross-reactivity suggests shared epitopes between M protein and phosphorylase, potentially impacting autoimmune responses.
Area of Science:
- Immunology
- Biochemistry
- Molecular Biology
Background:
- Streptococcal M proteins are key virulence factors.
- Autoimmunity can arise from molecular mimicry between microbial antigens and host proteins.
- Phosphorylases are enzymes involved in glycogen metabolism.
Purpose of the Study:
- To investigate immunological cross-reactivity between streptococcal M5 protein and rabbit muscle phosphorylases.
- To identify shared epitopes between M protein and phosphorylase.
- To assess the functional impact of this cross-reactivity on enzyme activity.
Main Methods:
- Enzyme-linked immunosorbent assay (ELISA) to screen for cross-reactivity.
- ELISA inhibition studies to confirm epitope sharing.
- Western blot (immunoblot) analysis to identify reacting protein subunits.
- Functional assays to measure enzyme inhibition by antibodies.
Main Results:
- Three of ten rabbit anti-M5 antisera showed significant cross-reactivity with phosphorylase a and b.
- Cross-reactive antibodies were inhibited by M5, phosphorylase b, M6, and M19, indicating shared epitopes across M protein serotypes.
- Anti-M5 antibodies recognized the subunit of phosphorylase b and partially inhibited phosphorylase a activity (up to 75%).
- Enzyme activity was restored upon pre-incubation of antibodies with M5 protein.
Conclusions:
- Shared epitopes exist between type 5 streptococcal M protein and muscle phosphorylases.
- These findings suggest a potential mechanism for autoimmune reactions in streptococcal infections.
- The cross-reactivity has functional consequences, including enzyme inhibition.