Structure-activity relationships within the N-terminal short consensus repeats (SCR) of human CR1 (C3b/C4b receptor, CD35): SCR 3 plays a critical role in inhibition of the classical and alternative pathways of complement activation

Genes coding for between one and four short consensus repeats (SCR) of the N-terminal region of human complement receptor 1 (CR1) were synthesized fro m oligonucleotides and those encoding SCR(1-2), SCR(1-3), SCR(1-4), SCR3 an d SCR(3-4) were expressed as inclusion bodies in Escherichia coli. Followin g solubilization in urea, the proteins were partially purified and refolded and the activity of each protein was assessed in both classical and altern ative pathway complement assays. All fragments showed a varying degree of a ctivity with the general order being SCR(1-3) drop SCR(1 -4)> SCR(1-2). Add ition of SCR3 to SCR(1-2) significantly improved potency, whereas the addit ion of SCR4 conferred no additional benefit. This observation, coupled with the ability of the single-domain SCR3 to inhibit classical pathway mediate d lysis with an IH50% (inhibition of hemolysis by 50 %) of 4.8 mu M, demons trates that SCR3 provides key binding interactions with activated complemen t components. SCR(1-3) was able to inhibit both classical and alternative p athways of complement activation, showing that the N-terminal SCR of CR1 re tain the ability to interact with C3b. Assays for CR1-like cofactor activit y for factor I using C4b-like C4 or C3b-like C3 as substrates showed that S CR(1-3) possessed such cofactor activity and that C4b-like C4 was a better substrate. When compared to full-length (30 SCR) soluble CRI (sCR1), SCR(1- 3) was significantly less potent in accord with a model involving multi-val ent binding of C3b/C4b to CR1.