MITOGENICITY OF M5 PROTEIN EXTRACTED FROM STREPTOCOCCUS-PYOGENES CELLS IS DUE TO STREPTOCOCCAL-PYROGENIC EXOTOXIN-C AND MITOGENIC FACTOR MF

M proteins of Streptococcus pyogenes are virulence factors which imped e phagocytosis, bind to many plasma proteins, and induce formation of cross-reactive autoimmune antibodies. Recently, it has been reported t hat some M proteins, extracted with pepsin from streptococci (pep M), are superantigens. One of these, pep M5, was investigated in detail an d was shown to Stimulate human T cells bearing V beta 2, V beta 4, and V beta 8. In the present study, we extracted and purified M5 protein by different biochemical methods from two M type 5 group A streptococc al strains. The crude extracts were fractionated by affinity chromatog raphy and ion-exchange chromatography. All fractions were tested in pa rallel for M protein by immunoblotting and for T-cell-stimulating acti vity. Although several crude preparations of M5 protein were associate d with mitogenicity for V beta 2 and V beta 8 T cells, the M5 proteins , irrespective of the extraction method, could be purified to the exte nt that they were no longer mitogenic. The mitogenic activity was not destroyed during the purification procedures but was found in fraction s separated from M protein. In these fractions, streptococcal pyrogeni c exotoxin C and mitogenic factor MF could be detected by protein blot ting and enzyme-linked immunosorbent assay. Moreover, anti-M protein s era did not inhibit the mitogenic activity of crude extracts, but anti sera which contained anti streptococcal pyrogenic exotoxin C antibodie s showed inhibition. The inability or M5 protein to stimulate T cells was confirmed with recombinant pep M5 produced in Escherichia coli. Ou r data strongly suggest that the mitogenic activity in M protein prepa rations is caused by traces of streptococcal superantigens different f rom M protein.