Biosynthesis of the internal thioester bond of the third component of complement

C3-translational product, which was synthesized with rabbit liver mRNA in a reticulocyte lysate protein-synthesizing system, did not react with [14C]methylamine, indicating the lack of an internal thioester bond. Instead, the C3-translational product reacted with iodo[1-14C]acetamide, as determined by gel electrophoresis in the presence of sodium dodecyl sulfate after immunoprecipitation of the product, indicating the presence of a reactive thiol group. When the C3-translational product was treated with rabbit liver homogenate, the product acquired reactivity with [14C]methylamine and lost the reactivity with iodo[1-14C]acetamide. Thus, the liver homogenate seemed to contain a factor (or factors) required for the formation of an internal thioester bond. The factor was partially purified from the liver homogenate by ammonium sulfate precipitation and ion-exchange chromatography on DEAE-cellulose.