Amino acid sequence of human D of the alternative complement pathway

Abstract

The primary structure of human D, the serine protease activating the C3 [complement component 3] convertase of the alternative complement pathway, was deduced by sequencing peptides derived from various chemical (CNBr [cyanogen bromide] and o-iodosobenzoic acid) and enzymatic (trypsin, lysine protease, Staphylococcus aureus V8 protease and chymotrypsin) cleavages. Carboxypeptidase A was also used to confirm the COOH-terminal sequence. The peptides were purified by high-pressure liquid chromatography. The proposed sequence of human D contains 222 amino acids and has a calculated MW of 23,748. It exhibits a high degree of homology with other serine proteases, especially around the NH2-terminus as well as the 3 residues corresponding to the active-site His-57, Asp-102 and Ser-195 (chymotrypsinogen numbering). This sequence homology is highest (40%) with plasmin, intermediate (35%) with pancreatic serine proteases, such as elastase, trypsin, chymotrypsin and kallikrein and least (30%) with the serum enzymes thrombin and factor X. D exhibits only minimal amino acid homology with the other sequenced complement serine proteases, Clr (25%) and Bb (20%). The substitution of a basic lysine for a neutral amino acid 3 residues NH2-terminal to the active-site serine as well as a small serine residue for a bulky aromatic amino acid at position 215 (chymotrypsinogen numbering) in the binding pocket may be important in determining the exquisite substrate specificity of D. The presence of His-40 which interacts with Asp-194 (chymotrypsinogen numbering) to stabilize other serine protease zymogens (Freer, S. T., Kraut, J., Robertus, J. D., Wright, H. T., and Xuong, N. H., 1970) argues in favor of such a D precursor molecule.