De novo coassembly of functional chlorophyll- and carotenoid-protein complexes in Saccharomyces cerevisiae

Chlorophylls are essential for photosynthesis, yet their heterologous production is a major challenge in synthetic biology due to pathway complexity and the diversion of resources away from essential cofactor biosynthesis. Here, we reconstruct the complete chlorophyll biosynthetic pathway in Saccharomyces cerevisiae. By integrating 13 heterologous genes and deleting 3 endogenous genes, we engineered a yeast chassis capable of producing chlorophyll a at titers of 26 micrograms per gram of dry cell weight. This platform was further expanded with a ketocarotenoid biosynthetic module to produce echinenone and canthaxanthin, enabling the in vivo coassembly of two distinct pigment-protein complexes: the plant-derived water-soluble chlorophyll protein and the cyanobacterial orange carotenoid protein (Syn_OCP). Spectroscopic and biochemical analyses confirm that these yeast-assembled complexes retain native spectral and functional signatures, including Syn_OCP photoactivity, demonstrating high-fidelity pigment binding. Our work establishes a versatile eukaryotic platform for producing photosynthetic complexes, which could help open avenues for studying their assembly outside plants and for engineering light-driven metabolism in nonphotosynthetic eukaryotes.