The Physiological Breakdown of C3b

Abstract The requirements for and products of C3b inactivator (KAF) action on C3b have been studied. C3 was purified from human serum. Final traces of contaminating proteins were removed by passage through a Sepharose “anti-impurity antiserum” column. C3b was generated by using insolubilized cobra venom factor convertase, and separated from C3a by chromatography on hydroxyl-apatite. The KAF-dependent cleavage of fluid phase C3b was found to have an absolute requirement for β1H (or possibly a contaminant of β1H preparations); neither KAF nor β1H alone had any discernable proteolytic action on C3b. The initial pattern of cleavage was found to be the same with purified KAF and β1H or with serum (as a source of these factors). The α-chain of C3b was cleaved into two polypeptide chains, an α1a chain of about 68K and an α1b chain of about 46K. The α1b chain was rapidly ‘trimmed’ to about 43K (α1b1). All of these three α-chain fragments were still covalently bonded to the β-chain. On antigenic analysis at this stage there was still a single component—presumably C3bi. More prolonged incubation with the β1H lead to cleavage of the α1a polypeptide, yielding finally a product of 29K (α1a2) no longer covalently bound to the β-chain. This is presumably C3d.