Recombinant C345C and factor I modules of complement components C5 and C7 inhibit C7 incorporation into the complement membrane attack complex.
Complement component C5 binds to components C6 and C7 in reversible reactions that are distinct from the essentially nonreversible associations that form during assembly of the complement membrane attack complex (MAC). We previously reported that the approximately 150-aa residue C345C domain (also known as NTR) of C5 mediates these reversible reactions, and that the corresponding recombinant module (rC5-C345C) binds directly to the tandem pair of approximately 75-residue factor I modules from C7 (C7-FIMs). We suggested from these and other observations that binding of the C345C module of C5 to the FIMs of C7, but not C6, is also essential for MAC assembly itself. The present report describes a novel method for assembling a complex that appears to closely resemble the MAC on the sensor chip of a surface plasmon resonance instrument using the complement-reactive lysis mechanism. This method provides the ability to monitor individually the incorporation of C7, C8, and C9 into the complex. Using this method, we found that C7 binds to surface-bound C5b,6 with a K(d) of approximately 3 pM, and that micromolar concentrations of either rC5-C345C or rC7-FIMs inhibit this early step in MAC formation. We also found that similar concentrations of either module inhibited complement-mediated erythrocyte lysis by both the reactive lysis and classical pathway mechanisms. These results demonstrate that the interaction between the C345C domain of C5 and the FIMs of C7, which mediates reversible binding of C5 to C7 in solution, also plays an essential role in MAC formation and complement lytic activity.
Thai
Chuong-Thu
CT
Torrey Pines Institute for Molecular Studies, San Diego, CA 92121, USA.
Ogata
Ronald T
RT
D020816:Amino Acid Motifs; D000818:Animals; D001667:Binding, Competitive; D003182:Complement C5 / Q000378:metabolism / Q000502:physiology*; D050776:Complement C5b; D003184:Complement C7 / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000502:physiology*; D017244:Complement Factor I / Q000502:physiology*; D003169:Complement Inactivator Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology; D015938:Complement Membrane Attack Complex / Q000037:antagonists & inhibitors* / Q000378:metabolism; D003171:Complement Pathway, Classical / Q000276:immunology; D003545:Cysteine; D006461:Hemolysis / Q000276:immunology; D006801:Humans; D007700:Kinetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D012756:Sheep; D020349:Surface Plasmon Resonance / Q000295:instrumentation / Q000379:methods; D013499:Surface Properties
D003182:Complement C5; D003184:Complement C7; D003169:Complement Inactivator Proteins; D015938:Complement Membrane Attack Complex; D011994:Recombinant Proteins; D050776:Complement C5b; D017244:Complement Factor I; D003545:Cysteine