Fitness of Taxadiene Biosynthetic Modules with Different S.cerevisiae Chassis

Abstract

In the research works of constructing taxadiene aritficaial synthetic cell,S.cerevisiae is a commonly used chassis.The highest taxadiene yield in S.cerevisiae that has been reported was 8.7 mg/L.In this work,the fitness of different taxadiene biosynthetic modules with different S.cerevisiae chassis was studied to elevate the taxadiene yield to a higher level.We chose the S.cerevisiae strains BY4742 and the single knockout strains of BY4742 as chassis,constructed the multi-plasmids taxadiene biosynthetic module by optimizing the mevalonic acid( MVA) pathway with tHMGR,importing the synthetic genes GGPPSsa and tTS,up-regulating and co-expressing the genes BTS1 and ERG20 which are correlated with the synthesis of geranylgeranyl diphosphate( GGPP).Then we redesigned and reconstructed the module inserted into a single CEN plasmid by the method of DNA assembler.Through fitting the different modules with different chassis,we acquired strains with diffe-rent taxadiene yields,the highest yield was 74.84 mg/L in the strain with the single-plasmid module and the chassis of YNL280C.The results indicated that the single-plasmid taxadiene biosynthetic module was more stable in the chassis than the multi-plasmids one,and the regulation of the pathways correlated with the MVA pathway will also influent the taxadiene yield.