Surface loops adjacent to the cation-binding site of the complement factor B von Willebrand factor type A module determine C3b binding specificity.
The interaction of factor B with C3b deposited on the surface of pathogens is the first step in the activation of the alternative complement pathway. The role of the von Willebrand factor type A (VWFA) module of factor B in this interaction has been investigated by generating three chimeras, Ch1-Ch3, in which surface loops of the VWFA module flanking the cation-binding residues were replaced by the corresponding sequences of C2, a factor B-like molecule which does not bind C3b. The location of the three loops was inferred from a homology model based on the structure of the integrin alphaM VWFA module [Ch1, betaA-alpha1 loop: Ch2, alpha3-alpha4 loop; and Ch3, betaD-alpha5 loop; Lee, J.-O., et al. (1995b) Cell 80, 631-638]. The function of the chimeras was studied by means of hemolytic assays and assays of the individual steps of the alternative complement pathway, i.e., binding to the C3b analogue cobra venom factor and factor D cleavage. These experiments showed that Ch1 and Ch3 define regions that are involved in C3b binding whereas Ch2 does not appear to be involved in binding specificity. The inability of Ch1 to register the enhancement of cobra venom factor binding normally seen after factor D cleavage suggested that the betaA-alpha1 loop mediates the conformational regulation of ligand binding affinity. Homology modeling of the chimeras has been used to visualize the surface structures which potentially define the C3b binding site.
Tuckwell
D S
DS
Wellcome Trust Centre for Cell-Matrix Research, School of Biological Sciences, University of Manchester, U.K.
Xu
Y
Y
Newham
P
P
Humphries
M J
MJ
Volanakis
J E
JE
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002412:Cations; D002417:Cattle; D002645:Chickens; D003179:Complement C3b / Q000737:chemistry / Q000378:metabolism*; D011415:Complement Factor B / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004546:Elapid Venoms / Q000378:metabolism; D004563:Electrochemistry; D006461:Hemolysis; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins; D017385:Sequence Homology; D014841:von Willebrand Factor / Q000737:chemistry* / Q000378:metabolism
D002412:Cations; D004546:Elapid Venoms; D011993:Recombinant Fusion Proteins; C016310:cobra venom factor; D014841:von Willebrand Factor; D003179:Complement C3b; D011415:Complement Factor B