IDENTIFICATION OF A HEPARIN-BINDING DOMAIN IN THE 7TH SHORT CONSENSUSREPEAT OF COMPLEMENT FACTOR H-1

Surface polyanions such as sialic acid and heparin are thought to enha nce the binding of complement factor H (fH) to C3b deposited on partic les and cell surfaces, thereby reducing complement activation, fH cont ains 20 short consensus repeat (SCR) domains, and it has been proposed that SCR 13 contains a heparin binding site, We used recombinant prot eins to determine the heparin binding site on fH, Full-length fH (H20) and truncated and SCR deletion mutant proteins were cloned and expres sed in Chinese hamster ovary cells, Supernatants were applied to hepar in-agarose affinity columns to determine their binding and elution pro files, Deletion of SCR 13 from H20 did not prevent heparin binding nor alter its salt elution profile, indicating that SCR 13 does not conta in an essential heparin binding site, We found that SCR 7 contains a h eparin binding site, as SCRs 1 through 7 were the smallest truncated p roteins to bind heparin (89 +/- 3%), Furthermore, deletion of SCR 7 fr om a protein containing SCRs 1 through 9 reduced heparin binding, wher eas deletion of SCR 6 did not (17 +/- 13 vs 81 +/- 13%; p = 0.02), It is likely that other heparin binding sites exist within SCRs 10 throug h 20; an SCR 7 deletion mutant of H20 eluted earlier than H20, but sti ll showed >99% binding to immobilized heparin, SCR 13 does not contain such a site because a double deletion of SCRs 7 and 13 from H20 showe d >97% heparin binding and had an elution profile smilar to that of a single deletion of SCR 7.