CCP1-4 of the C4b-binding protein alpha-chain are required for factor I mediated cleavage of complement factor C3b.
C4b-binding protein (C4BP) is a potent regulator of the complement system because it strongly inhibits the classical pathway of complement. Furthermore, C4BP serves as a cofactor to factor I (FI) in the cleavage of fluid phase C3b and can, therefore, influence the alternative pathway of complement. The major form of C4BP in plasma consists of seven identical alpha-chains and one beta-chain. Both types of subunits are composed of complement control protein (CCP) domains, eight such domains make up one alpha-chain. To elucidate the structural requirements for the interaction between C3b and the alpha-chain, nineteen recombinant C4BP variants were used: six truncated monomeric variants, nine polymeric variants in which individual CCPs were deleted, and finally four variants in which double alanine residues were introduced between CCPs. We found that C4BP requires all four N-terminal CCPs of the alpha-chain, with CCP2 and 3 being the most important, to act as a cofactor in the cleavage of C3b. Also, a cluster of positively charged amino acids on the interface between CCP1 and 2 is involved in the binding. Compared to the interaction with C4b, we conclude that binding of C3b to C4BP requires larger molecular surface on C4BP. We found that C4BP was able to act as cofactor in degradation of surface bound C3b and to accelerate decay of alternative C3-convertase. However, in both cases 1,000-fold molar excess of C4BP over factor H (FH), well known inhibitor of the alternative pathway, was required to obtain the same effect.
Blom
Anna M
AM
The Wallenberg Laboratory, Department of Clinical Chemistry, Lund University, University Hospital Malmö, S-205 02 Malmö, Sweden. anna.blom@klkemi.mas.lu.se
Kask
Lena
L
Dahlbäck
Björn
B
D019943:Amino Acid Substitution; D000818:Animals; D003167:Complement Activation; D050577:Complement C3-C5 Convertases / Q000378:metabolism; D003179:Complement C3b / Q000378:metabolism*; D017242:Complement Factor H / Q000378:metabolism; D017244:Complement Factor I / Q000378:metabolism*; D003169:Complement Inactivator Proteins / Q000235:genetics / Q000378:metabolism*; D006023:Glycoproteins; D009154:Mutation; D017434:Protein Structure, Tertiary / Q000502:physiology; D012756:Sheep
D003169:Complement Inactivator Proteins; D006023:Glycoproteins; D003179:Complement C3b; D017242:Complement Factor H; D050577:Complement C3-C5 Convertases; D017244:Complement Factor I