Structural basis of the binding specificity of the thioester-containing proteins, C4, C3 and alpha-2-macroglobulin.

We have previously noted a large difference in the specificity of the covalent binding reaction of human C4-A and C4-B. Here we report data on three other thioester-containing proteins. Human C3 is unreactive with glycine but its reactivity with glycerol (k'/ko = 23.0 M-1) is similar to that of human C4-B (k'/ko = 15.5 M-1). Human alpha 2-macroglobulin reacts with glycine (k'/ko = 206 M-1) in a manner similar to C4-B (k'/ko = 119 M-1) but its reactivity with glycerol (k'/ko = 1.2 M-1) is C4-A like (k'/ko = 1.3 M-1). Mouse C4 is C4-B like in its reaction with both glycine (k'/ko = 136 M-1) and glycerol (k'/ko = 26.0 M-1). Of these proteins, only C4-A shows a very high rate of reaction with glycine (k'/ko = 13,400 M-1). The comparison of the primary structures of these proteins has allowed us to propose the Leu Asp:Ile His substitutions at positions 1105 and 1106 in the human pro-C4 molecule as the residues largely responsible for the binding specificities of these proteins. The Leu:Ile change would not markedly affect the reactivity of these proteins, but may be necessary for allosteric reasons. The Asp in C4-A and His in C4-B seem likely to be the major specificity-defining residues.