Substitution of a single amino acid (aspartic acid for histidine) converts the functional activity of human complement C4B to C4A.
The C4B isotype of the fourth component of human complement (C4) displays 3- to 4-fold greater hemolytic activity than does its other isotype C4A. This correlates with differences in their covalent binding efficiencies to erythrocytes coated with antibody and complement C1. C4A binds to a greater extent when C1 is on IgG immune aggregates. The differences in covalent binding properties correlate only with amino acid changes between residues 1101 and 1106 (pro-C4 numbering)--namely, Pro-1101, Cys-1102, Leu-1105, and Asp-1106 in C4A and Leu-1101, Ser-1102, Ile-1105, and His-1106 in C4B, which are located in the C4d region of the alpha chain. To more precisely identify the residues that are important for the functional differences, C4A-C4B hybrid proteins were constructed by using recombinant DNA techniques. Comparison of these by hemolytic assay and binding to IgG aggregates showed that the single substitution of aspartic acid for histidine at position 1106 largely accounted for the change in functional activity and nature of the chemical bond formed (ester vs. amide). Surprisingly, substitution of a neutral residue, alanine, for histidine at position 1106 resulted in an increase in binding to immune aggregates without subsequent reduction in the hemolytic activity. This result strongly suggests that position 1106 is not "catalytic" as previously proposed but interacts sterically/electrostatically with potential acceptor sites and serves to "select" binding sites on potential acceptor molecules.
Department of Pathology, Harvard Medical School, Boston, MA.
M C
MC
Carroll
D M
DM
Fathallah
L
L
Bergamaschini
E M
EM
Alicot
D E
DE
Isenman
D000818:Animals; D001224:Aspartic Acid; D001483:Base Sequence; D002460:Cell Line; D015934:Complement C4a / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D015935:Complement C4b / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D005822:Genetic Vectors; D006461:Hemolysis; D006639:Histidine; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000494:pharmacology; D014162:Transfection
D015345:Oligonucleotide Probes; D011994:Recombinant Proteins; D001224:Aspartic Acid; D006639:Histidine; D015934:Complement C4a; D015935:Complement C4b