Identity of the residues responsible for the species-restricted complement inhibitory function of human CD59.
The membrane-anchored glycoprotein CD59 inhibits assembly of the C5b-9 membrane attack complex (MAC) of human complement. This inhibitory function of CD59 is markedly selective for MAC assembled from human complement components C8 and C9, and CD59 shows little inhibitory function toward MAC assembled from rabbit and many other non-primate species. We have used this species selectivity of CD59 to identify the residues regulating its complement inhibitory function: cDNA of rabbit CD59 was cloned and used to express human/rabbit CD59 chimeras in murine SV-T2 cells. Plasma membrane expression of each CD59 chimera was quantified by use of a 5'-TAG peptide epitope, and each construct was tested for its ability to inhibit assembly of functional MAC from human versus rabbit C8 and C9. These experiments revealed that the species selectivity of CD59 is entirely determined by sequence contained between residues 42 and 58 of the human CD59 polypeptide, whereas chimeric substitution outside this peptide segment has little effect on the MAC inhibitory function of CD59. Substitution of human CD59 residues 42-58 into rabbit CD59 resulted in a molecule that was functionally indistinguishable from native human CD59, whereas the complementary construct (corresponding residues of rabbit CD59 substituted into human CD59) was functionally indistinguishable from rabbit CD59. Based on the solved solution structure of CD59, these data suggest that selectivity for human C8 and C9 resides in a cluster of closely spaced side chains on the surface of CD59 contributed by His44, Asn48, Asp49, Thr51, Thr52, Arg55, and Glu58 of the polypeptide.
Zhao
X J
XJ
Blood Research Institute, the Blood Center, Milwaukee, Wisconsin 53201-2178, USA.
Zhao
J
J
Zhou
Q
Q
Sims
P J
PJ
D000595:Amino Acid Sequence; D000818:Animals; D018957:CD59 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002645:Chickens; D003001:Cloning, Molecular; D003169:Complement Inactivator Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary; D006801:Humans; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D018957:CD59 Antigens; D003169:Complement Inactivator Proteins; D018076:DNA, Complementary; D011994:Recombinant Proteins