Construction and expression in the yeast Pichia pastoris of functionally active soluble forms of the human costimulatory molecules B7-1 and B7-2 and the B7 counter-receptor CTLA-4

Abstract

We have generated soluble recombinant forms of the costimulatory molecules B7-1 and B7-2, and their counter-receptor CTLA-4 using a yeast Pichia pastoris expression system. Fragments comprising the extracellular domains of human B7-1, B7-2, and CTLA-4 molecules were expressed at high levels and could be purified from culture supernatants following a simple one-step purification protocol. The recombinant proteins retained their functionality and specific binding to their natural counterparts could be demonstrated by FACS analysis. In T cell proliferation assays costimulatory activity of immobilized B7-1 and B7-2 proteins in the presence of an anti-CD3 antibody was observed with the B7-1 protein being more potent than B7-2.