C9 hemolytic activity of the soluble C5b-9 complex of guinea pig complement, analogous to human SC5b-9
Kinoshita, T.; Hong, K.; Inoue, K.
Journal of Immunology 123(5): 1989-1995
1979
ISSN/ISBN: 0022-1767 PMID: 114583 Document Number: 152685
When guinea pig serum was treated with zymosan, hemolytic activity of C9 [complement component 9] appeared in a macromolecular complex with a MW of about 1,000,000 and free C9. The complex was isolated by sequential gel filtrations and sucrose density gradient centrifugation. On immunodiffusion analysis C5 and C9 incorporated into the complex showed loss of some antigenic determinants of the respective precursor proteins. Immunoelectrophoretic analysis revealed that the complex moved faster than free C9. Preliminary estimation of the subunit composition of the complex by sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis showed that it contained similar proteins to those found in the human SC5b-9 complex. The complex seemed to contain 1 molecule each of C5b, C6, C7, C8 and protein corresponding to human S protein and 3 molecules of C9. The C9 hemolytic activity of the complex was very similar to that of free C9 dose dependently and kinetically. It was neutralized with antibody [Ab] to free C9 in a first order fashion, but the Ab to C5 did not neutralize hemolytic activity in spite of its ability to precipitate the complex. The complex showed C9 hemolytic activity and slight C7 activity. No other hemolytic activity was found when assayed with all the intermediate cells along the classical pathway or when assayed as any other components of C. The human counterpart, SC5b-9, showed no C9 hemolytic activity but competitively inhibited the hemolytic activity of free C9.