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Sophorolipids (SLs), an important biosurfactant produced by  S. bombicola , were one of the most potential substitutes for chemical surfactants. Few reports on the transcriptional regulation of SLs synthesis and the engineered strains with high-yield SLs were available. In this study, a Rim9-like protein (Rlp) and three transcription factors ( ztf1 ,  leu3 ,  gcl ) were mined and analyzed, and a progressive enhancement of SLs production was achieved through cumulative knockouts of three genes. The sophorolipid production of  ΔrlpΔleu3Δztf1  reached 97.44 g/L, increased by 50.51% than that of the wild-type strain. Compared with the wild-type strain, the flow of glucose to SLs synthesis pathways was increased, and the synthesis of branched-chain amino acids was reduced in  ΔrlpΔleu3Δztf1 . The amount of UDP-glucose, the substrate for two glycosyltransferases, also increased, and the expression level of the key genes  sble  and  UGPase  for SLs synthesis increased by 2.2 times, respectively. The multiple-gene knockout strategy was proved to be highly effective to construct the engineered strain with high-yield SLs production, and this strain was a superior strain for industrial fermentation of SLs and reduced SLs production costs.

frontiers in bioengineering and biotechnology

A Cumulative Effect by Multiple-Gene Knockout Strategy Leads to a Significant Increase in the Production of Sophorolipids in Starmerella Bombicola CGMCC 1576