Isolation and Purification of C9 and Vitronectin from Goat Plasma

Abstract

Background: Vitronectin (Vn) is a multifunctional protein of blood and extracellular matrix. It binds to complement C9, which is involved in the terminal step of cytolysis including those of microbes. It hypothesized that Vn may bind to pore farming complement protein i.e C9 to modulate innate immunity and involved in immune evasion mechanism of bacterial pathogens like staphylococcus aureus. Methods: The procedure allows the rapid, large-scale isolation of pure and haemolytically active Vn and C9 proteins from goat plasma by suitable ion-exchange and affinity chromatographic techniques. Result: Vitronectin (Vn), a multifunctional protein of blood and the extracellular matrix has a mol.wt.of 78 kDa. Goat C9, the last component of complement, was purified in good yield by a combination of salt fractionation and ion-exchange chromatography. Approximately 1mg of protein can be obtained from 1 litre of the serum. The C9 was obtained has a mol.wt.of 66 kDa, determined by SDS/polyacrylamide-gel electrophoresis. No impurities were detected on gel electrophoresis and the Vn and C9 were confirmed by western blotting.