Identification of intermolecular bonds between human factor B and Cobra Venom Factor important for C3 convertase stability

Abstract

Cobra venom factor (CVF) is the complement-activating protein in cobra venom. CVF is a structural and functional analog of complement component C3. CVF, like C3b, forms a convertase with factor B. This bimolecular complex CVF, Bb is an enzyme that cleaves C3 and C5. However, CVF, Bb exhibits significantly different functional properties from C3b,Bb. Whereas both, CVF, Bb and C3b, Bb exhibit spontaneous decay-dissociation into the respective subunits, thereby eliminating the enzymatic activity, the CVF, Bb convertase is physico-chemically far more stable, decaying with a half-life that is more than two orders of magnitude slower than that of C3b,Bb. In addition, CVF, Bb is completely resistant to inactivation by Factors H and I. These two properties of CVF, Bb allow continuous activation of C3 and C5, and complement depletion in serum. In order to understand the structural basis for the physico-chemical stability of CVF,Bb, we have created recombinant hybrid proteins of CVF and human C3, based on structural differences between CVF and human C3b in the C-terminal C345C domain. Here we describe three human C3/CVF hybrid proteins which differ in only one, two, or five amino acid residues from earlier described hybrid proteins. In all three cases, the hybrid proteins containing CVF residues form more stable convertases, and exhibit stronger complement-depletion activity than hybrid proteins with human C3 residues. Three bonds between CVF residues and Factor Bb residues could be identified by crystallographic modeling that contribute to the greater stability of the convertases.

Keywords: C3 convertase; Cobra (Naja sp.); Cobra venom factor; Complement; Complement component C3; Hybrid proteins.