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|Title:||Identification of a heparin binding domain in the seventh short consensus repeat of complement factor H|
|Citation:||Journal of Immunology, 1996; 157(12):5422-5427|
|Publisher:||American Association of Immunologists|
|Timothy K. Blackmore, Tania A. Sadlon, Helena M. Ward, Douglas M. Lublin, and David L. Gordon|
|Abstract:||Surface polyanions such as sialic acid and heparin are thought to enhance the binding of complement factor H (fH) to C3b deposited on particles and cell surfaces, thereby reducing complement activation. fH contains 20 short consensus repeat (SCR) domains, and it has been proposed that SCR 13 contains a heparin binding site. We used recombinant proteins to determine the heparin binding site on fH. Full-length fH (H20) and truncated and SCR deletion mutant proteins were cloned and expressed in Chinese hamster ovary cells. Supernatants were applied to heparin-agarose affinity columns to determine their binding and elution profiles. Deletion of SCR 13 from H20 did not prevent heparin binding nor alter its salt elution profile, indicating that SCR 13 does not contain an essential heparin binding site. We found that SCR 7 contains a heparin binding site, as SCRs 1 through 7 were the smallest truncated proteins to bind heparin (89 ± 3%). Furthermore, deletion of SCR 7 from a protein containing SCRs 1 through 9 reduced heparin binding, whereas 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 through 20; an SCR 7 deletion mutant of H20 eluted earlier than H20, but still 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 showed >97% heparin binding and had an elution profile smilar to that of a single deletion of SCR 7.|
|Keywords:||CHO Cells; Animals; Humans; Heparin; Complement Factor H; Recombinant Proteins; Cloning, Molecular; Sequence Deletion; Binding Sites; Consensus Sequence; Protein Binding; Structure-Activity Relationship; Cricetinae|
|Rights:||Copyright © 1996 by American Association of Immunologists|
|Appears in Collections:||Medical Education Unit publications|
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