The molecular basis of the low hemolytic activity of C4 molecules from low-C4 mice with IgM-coated erythrocytes.
This study investigated the origin of the different hemolytic activity of two allotypes of murine C4, C4H (C4-high) and C4L (C4-low) in the presence of IgM-coated erythrocytes. C4H displayed a threefold higher hemolytic titer (expressed in hemolytic units/microgram protein) than C4L. No difference was found between c4H and C4L either in stability at 37 degrees C at different pH values and in the rate of C4H and C4L hydrolysis by activated Cl. The major functional difference was found in the covalent binding capacity to IgM-coated erythrocytes, with the amount of C4H bound being about threefold higher than that of C4L. A marked difference in the reactivity of the C4b fragment of C4H and C4L toward amino and hydroxyl groups was detected. Using glycine and glucose to competitively inhibit the binding of C4H and C4L, it was observed that C4L selects preferentially for the amino groups of glycine, whereas C4H reacted preferentially with the hydroxyl groups of glucose. Furthermore, the chemical modification of the red cells amino groups by ethylacetimidate, had no appreciable effect on the binding of C4H, but reduced that of C4L. Southern blot hybridization experiments using Hind III-digested liver DNA and a cDNA probe derived from the 5' end of the mRNA (clone pAT-A) indicated that the C4H allele was associated with a 5-kb Hind III fragment, whereas the C4L allele was associated with a 15-kb Hind III fragment. The possibility that a structural difference between C4H and C4L is responsible for their different reactivities is discussed.
Dieli
F
F
Institute of General Pathology, University of Palermo, Italy.
Sireci
G
G
Lio
D
D
Salerno
A
A
D000483:Alleles; D000818:Animals; D015139:Blotting, Southern; D003167:Complement Activation; D003176:Complement C3 / Q000378:metabolism; D003181:Complement C4 / Q000502:physiology*; D003182:Complement C5 / Q000378:metabolism; D004912:Erythrocytes / Q000276:immunology; D005796:Genes; D006461:Hemolysis / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007096:Imidoesters / Q000494:pharmacology; D008744:Methylamines / Q000494:pharmacology; D051379:Mice; D008815:Mice, Inbred Strains; D011485:Protein Binding
D003176:Complement C3; D003181:Complement C4; D003182:Complement C5; D007096:Imidoesters; D008744:Methylamines; C014539:ethyl acetimidate; C027451:methylamine