PRODUCTION OF GENETICALLY ENGINEERED LOW-AFFINITY HUMAN IMMUNOGLOBULIN RECEPTORS FCγRIIA (CD32A) AND FCγRIIIA (CD16A) EXTRACELLULAR DOMAINS IN PICHIA PASTORIS

Abstract: Genetic engineering technologies allow mass production of proteins for a variety of applications mainly in the biopharmaceutical industry. FcγRIIa (CD32a) and FcγRIIIa (CD16a) are antibodies cell surface receptors that mediate important immune functions. Fcγ receptors are currently the focus in developing novel therapeutic antibodies. Herein, we report the production at a high yields of genetically engineered soluble forms of the extracellular domain of the main genetic variants of CD16a (158F/V) (1.5mg/ml, 0.75mg/ml) and CD32a (131H/R) (5.8mg/ml, 0.9mg/ml) by the yeast Pichia pastoris. The Proteins were produced following induction using 1% methanol. SDS-PAGE and Western blot analysis of the protein purified on Ni+ column showed bands of ~45 kDa and ~25 kDa corresponding to CD16a and CD32a respectively. The production at a high yield of good quality recombinant Fcγ receptors in yeast is an important outcome because of their importance in the development of third generation therapeutic antibodies.

Keywords: FcγRIIa (CD32a), FcγRIIIa (CD16a), recombinant protein, Pichia pastoris, pPICZαA.