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

Jun Liu,Xinyu Zhang,Guoqin Zhao,Xin Song,Xiaoran Fang,Guodong Liu

doi:10.3389/fbioe.2022.818445

Jun Liu, Xinyu Zhang + Show 4 more

Open Access

https://doi.org/10.3389/fbioe.2022.818445

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Abstract

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.

Highlights

Surfactants are amphipathic compounds with both hydrophilic and hydrophobic moieties and have a wide range of applications
It is speculated that there is a homologous protein of rim9 in S. bombicola, which affects the synthesis of SLs by directly or indirectly affecting the expression of key genes for SLs synthesis
Through analyzing the results of qRT-PCR (Figure 5B), the expression levels of the key genes sble and UGPase for SLs synthesis in Δrlp were 2.1 and 3.9 times higher than those of the wild-type strain. These results indicated that deletion of rlp reduces the synthesis of polysaccharides from UDP-glucose and increases the high expression of the key enzyme genes for SLs synthesis to a certain extent

Summary

Introduction

Surfactants are amphipathic compounds with both hydrophilic and hydrophobic moieties and have a wide range of applications. Most surfactants in use today are petroleum-based and chemically synthesized. Such chemically-synthesized surfactants are difficult to degrade through the action of microorganisms (Santos et al, 2016), and cause significant ecological problems, in washing applications as these surfactants inevitably end up in the environment after use. Biosurfactants have several advantages over their chemically produced counterparts. Such advantages involve non-toxicity, biodegradability, bioavailability, biocompatibility, eco-friendliness, effectiveness and stability at extreme environments (active in broad range of pH, temperature and salt concentrations), and long storage time.

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