Corynebacterium glutamicum is an important strain for the industrial production of amino acids, but the fermentation of L-methionine has not been realized. The purpose of this study is to clarify the effect of reducing power NADPH on L-methionine synthesis. Site-directed mutagenesis of zwf and gnd genes in pentose phosphate pathway relieved feedback inhibition, increased NADPH supply by 151.8%, and increased L-methionine production by 28.3%; Heterologous expression of gapC gene to introduce NADP+ dependent glyceraldehyde-3-phosphate dehydrogenase increased NADPH supply by 75.0% and L-methionine production by 48.7%; Heterologous expression of pntAB gene to introduce membrane-integral nicotinamide nucleotide transhydrogenase increased NADPH by 89.2% and L-methionine production by 35.9%. Finally, the engineering strain YM6 with a high NADPH supply was constructed, which increased the NADPH supply by 348.2% and the L-methionine production by 64.1%. The analysis of metabolic flux showed that YM6 significantly increased the glycolytic flux, including the metabolic flux of metabolites such as glycosyldehyde-3-phosphate, dihydroxyacetate phosphate, 3-phosphoglycate and pyruvate, and the significant increase of L-methionine flux also confirmed the increase of its synthesis. This study provides a research basis for the systematic metabolic engineering construction of L-methionine high-yield engineering strains.

Sporosarcina aquimarina MS4 is a microecological preparation for overwintering Apostichopus japonicus, which has an immune regulation function, but its role in the nutritional regulation of A. japonicus is not clear. This study aimed to describe the effects of S. aquimarina MS4 on the growth, digestion, and body wall nutrition of A. japonicus through feeding experiments and to discuss the potential mechanism of S. aquimarina MS4 regulating gut function through the detection of gut microbiota and metabolites. After 60 days of culture, the growth performance of A. japonicus fed S. aquimarina MS4 (108 cfu/g) significantly improved, and the content of polysaccharide, leucine, phenylalanine, lysine, and docosahexaenoic acid in the body wall significantly increased. Gut microbiota analysis showed that although Proteobacteria, Verrucomicrobia, Firmicutes, and Bacteroidetes were the predominant phyla in all the sea cucumbers, Haloferula and Rubritalea showed significant difference between the group fed with or without S. aquimarina MS4. Metabolomics analysis showed that differential metabolites in the gut were mainly enriched in amino acid metabolism and lipid metabolism. The association analysis of differential metabolites and microbiota showed that the production of some differential metabolites was significantly related to differential microorganisms, which improved the understanding of the function of microorganisms and their roles in the gut of A. japonicus. This study reveals the life activities such as growth and metabolism of A. japonicus, and it provides support for the functional study of the gut microbiome of A. japonicus.