The low enoyl-acyl carrier protein reductase activity of FabI2 is responsible for the high unsaturated fatty acid composition in Sinorhizobium meliloti

Abstract

Abstract Background Sinorhizobium melilotiis recognized for its remarkable production of unsaturated fatty acids (UFAs). Prior investigations have demonstrated that S. meliloti predominantly utilizes the FabA–FabB pathway for UFAs, however, the mechanisms remain elusive. This study aimed to resolve the mechanisms of Sinorhizobium meliloti producing remarkable levels of UFAs. Methods The enoyl-acyl carrier protein (ACP) reductase (ENR) encoded genes were disrupted by suicide plasmid pK18mobsacB, then single-crossover and double-crossover. ENR proteins were purified in E.coli BL21(DE3) strains and ENR activities were tested by gel electrophoresis, NADH oxidation. The fatty acid composition were analyzed by gas chromatography-mass spectrometry (GC-MS) and thin-layer chromatography. Results Our results revealed that the heterogeneous expression fabI2 in an Escherichia coli fabI temperature-sensitive mutant leads to a substantial increase in unsaturated fatty acids production. Genetic analysis confirmed that fabI2 is an essential gene and cannot be disrupted in the S. meliloti genome. Intriguingly, we found that fabI2 was only functionally replaced by Enterococcus faecalis fabI, but not by S. meliloti fabI1, E. coli fabI, or Pseudomonas aeruginosa fabV. Moreover, we confirmed that the deletion of fabI1caused S. meliloti to produce more unsaturated fatty acids than the wild-type strain Rm1021. Conclusions In this study, we reported an enoyl-acyl carrier protein (ACP) reductase, encoded by the S. meliloti SMc00326 gene (fabI2), plays a key role in the production of unsaturated fatty. the FabI1, encoded by SMc00005, modulated the fatty acid compositionof S. meliloti. Collectively, these findings lay the foundation for proposing a model that elucidates the robust capacity of FabI2 in driving unsaturated fatty acid synthesis in S. meliloti.