Comparative Transcriptome Analysis of Key Reductase Genes Involved in the 1-Deoxynojirimycin Biosynthetic Pathway in Mulberry Leaves and Cloning, Prokaryotic Expression, and Functional Analysis of 1 and 2.

The alkaloid 1-deoxynojirimycin (DNJ) is the main bioactive ingredient in the hypoglycemic action of mulberry leaves ( L.). Our previous research clarified the upstream pathway from lysine to Δ1-piperideine in the biosynthesis of DNJ in mulberry leaves, but the pathway and related reductase genes from Δ1-piperideine to piperidine are still unclear. Here, a comparative transcriptome was used to analyze the transcriptome data of two samples (July and November) of mulberry leaves with significant differences in the content of DNJ and screen-related reductase genes. Results showed that expression levels of 1 and 2 were significantly and positively correlated with the content of DNJ ( < 0.05) in different seasons. 1 (GenBank accession no. MT989445) and 2 (GenBank accession no. MT989446) were successfully cloned and used for prokaryotic expression and functional analysis in vitro. 1 and 2 could catalyze the reaction of Δ1-piperideine with the coenzyme NADPH to generate piperidine. The kinetic parameters of 1 and 2 indicated that 2 had a higher binding ability to Δ1-piperideine than 1. This study provided insights into the biosynthesis of DNJ in mulberry leaves.