Abstract

Microalgae have become increasingly important in the biotech sector and are currently exploited for their natural products. In recent years efforts have also been directed towards establishing them as production platforms for recombinant proteins. I set out to investigate the feasibility of Chlamydomonas reinhardtii as a production platform for high-value products expressed in the chloroplast of the alga. A cytochrome P450 was introduced into the chloroplast of C. reinhardtii as a proof-of-concept study. The model enzyme CYP79A1 was successfully targeted into the chloroplast membranes and was found to be active. A bifunctional diterpene synthase, TPS4 , was also expressed in the chloroplast of C. reinhardtii. TPS4 could be purified to homogeneity and is the largest enzyme expressed in the chloroplast to date. The two transgenic strains expressing CYP79A1 and TPS4 were investigated for their ability to withstand industrial growth conditions. The cell wall deficient strains were successfully cultivated in 100 L photobioreactors using a mixotrophic growth regime. They reached dry weights of 0.3 g/L and the expression of CYP79A1 and TPS4 was detected over the entire growth period. Taken together these data suggest that C. reinhardtii could be an attractive platform for recombinant protein production. Two enzymes with biotechnological relevance were expressed in the chloroplast of the alga and the transgenic cell wall deficient strains were successfully cultivated on a semi-industrial scale.