Bioplastic Polyhydroxybutyrate production in the genetically engineered microalga Chlamydomonas reinhardtii UVM4

Abstract

Sustainability of production and consumption of plastic remains a pressing matter in our time. The non-degradable nature of conventional plastics poses a growing concern in terms of environmental health and even effects on human health and physiology. Bioplastics are becoming an increasingly popular alternative to conventional plastics. Some bioplastics such as polyhydroxybutyrate (PHB) has been observed to inhabit many of the same physiochemical properties of conventional plastics like high thermostability, reliable mechanical properties (strength and flexibility), low oxygen permeability and ultraviolet radiation (UV) resistance, while also being entirely biodegradable and biocompatible. Still, some challenges to widespread adoption and production of bioplastics are high costs, especially associated with feed stock expenses. Utilization of microalgae introduces an alternative to the traditionally used bacterial fermenters and a notable potential reduction in feed stock related costs. In this study, the biotechnological model microalga Chlamydomonas reinhardtii was utilized as a chassis for endogenous synthetic PHB production. Native bacterial genes phbB and phbC from Cupriavidus necator were successfully integrated into the host nuclear genome. Three different subcellular compartments was targeted to investigate optimum transgene expression and subsequent production of PHB. PHB was produced by the transgenic microalgae as ascertained by plate reader measurements. Integration of phbB and phbC was sufficient to facilitate PHB production, proving native presence of phbA in C.reinhartii. A significant difference (p < 0,05) was observed between the non-transformed control group UVM4 and the mitochondria and chloroplast targeted groups, but not with cytosol targeted strains in terms of PHB production. The chloroplast targeted strains showed the highest transgene expression by proxy of fluorescence (256) and also the highest synthetic production of PHB (0,23 μg/ml). Significant differences were also observed between target strains group, indicating potential optimal subcellular targets. The cytosolic strains illustrated the most visible presence of PHB, but starch and lipid presence trumped PHB visibility. For future enhancements of endogenous PHB production in C. reinhardtii, taking additional steps to further funnel metabolic flux towards PHB and steering it away from other biomolecules such as starches and lipids should be greatly encouraged.