Enhanced Rhamnolipid Production of Pseudomonas aeruginosa DN1 by Metabolic Engineering under Diverse Nutritional Factors

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Rhamnolipids are a very promising class of biosurfactants exhibiting properties of great interest in several industrial applications, but they are not widely used because of the low yield and the high cost of production. In this study, a metabolic engineering strategy was used to construct the engineered strains DNAB and DNC through introducing rhlAB and rhlC genes respectively to Pseudomonas aeruginosa wild-type strain DN1, as well as optimization of nutritional parameters such as carbon and nitrogen sources were assessed simultaneously, with the purpose of promoting the productivity of rhamnolipids. Both engineered strain DNAB and DNC had higher yield of rhamnolipids than the DN1 under the same conditions by means of increasing the copy number of rhlAB and rhlC genes, respectively. Of particular importance was olive oil and sodium nitrate as the optimal sole carbon and nitrogen source separately, engineered strain DNAB had the highest rhamnolipid yields 1.28-fold and 1.25-fold of the DN1, and engineered strain DNC had the highest rhamnolipid yields 1.36-fold and 1.43-fold of the DN1. The ideal C/N ratio was found to be 20 that increased specific rhamnolipid productivity to 19.5 g/L and 22.5 g/L of the engineered strains, a certain amount to 1.39-fold and 1.61-fold of DN1 strain respectively. Meanwhile, there was a difference of the identified rhamnolipids between wild-type and engineered strains by ESI-MS analysis, and Rha- Rha-C10 and Rha-Rha-C10-C10 were the most dominant structure of rhamnolipids produced by the engineered strains through altering the expression levels of RhlAB and RhlC.