Abstract
Background: β-carotene is a precursor of vitamin A and has great commercial value as an additive in foods and feeds. Many pathways not directly related to the β-carotene synthesis affect β-carotene production since the interactions among metabolic fluxes of cells confer a complex regulatory network. Engineered Y. lipolytica strain has excellent potential for β-carotene production as oleaginous yeast. Optimizing indirectly metabolic pathways in Y. lipolytica may offer a new strategy for making the β-carotene production achieve a commercially viable yield.Results: In this study, we found that the proper promotion of the multivesicular body (MVB) sorting pathway elevated the production of β-carotene by 1.58 fold when overexpressing one copy of the Did2 gene in Y. lipolytica. Through the measurement of ATP, NADPH, the mRNA, and protein level of key genes in the β-carotene synthesis pathway, the reason for β-carotene elevated was deuced that the protein level of the key enzymes (tHMG and CarA) was increased. When overexpressing two copies of the Did2 gene, the transcription level of the key genes was all elevated. However, the protein level of key enzymes in the β-carotene synthesis pathway was reduced compared with the overexpressing one copy of the Did2 gene, which resulted in reduced β-carotene content.Conclusion: This study suggests that the MVB sorting pathway is not responsible for sorting protein but has a crucial regulating effect on protein abundance in cells. Engineering the MVB sorting pathway could potentially increase the production of other high-value products. Moreover, manipulation of indirectly related metabolic pathways also is a critical strategy in synthetic biology research.
Studies in carotenogenesis. 1. General conditions governing β-carotene synthesis by the fungus Phycomyces blakesleeanus Burgeff
