HIGH-LEVEL EXPRESSION OF A FUNCTIONAL HUMAN-FIBRINOGEN GAMMA-CHAIN IN ESCHERICHIA-COLI

Human fibrinogen .gamma. chain has been expressed intact at high levels in Escherichia coli. The construction of the expression plasmid, p253, is described. Synthesis of the recombinant protein is isopropyl-.beta.-D-thiogalactopyranoside-dependent and is driven by the tac promoter. Western analysis of E. coli lysates demonstrates a novel protein of approx. 45 kDa which cross-reacts with antisera to human fibrinogen .gamma. chains. The protein is not soluble in common E. coli lysis buffers and becomes soluble in 6 M guanidine .cntdot. HCl or 6 M urea. Initial insolubility is due to interchain disulfide bond formation and to noncovalent interactions. Induced cells examined by phase-contrast microscopy contain dense inclusion bodies. A known function of the .gamma. chains of human fibrinogen is the clumping of Staphylococcus aureus Newman D2C cells [Hawiger et al., Biochemistry (1982) 1407-1413]. We demonstrate that suspensions of recombinant .gamma. chains retain the ability to clump cells from this strain of S. aureus.