Functional Analysis of 3-Dehydroquinate Dehydratase/Shikimate Dehydrogenases Involved in Shikimate Pathway in .

Polyphenols play an important role in the astringent taste of tea [ (L.)] infusions; catechins in phenolic compounds are beneficial to health. The biosynthesis of gallic acid (GA), a precursor for polyphenol synthesis, in tea plants remains unknown. It is well known that 3-dehydroquinate dehydratase/shikimate dehydrogenase (DQD/SDH) is a key enzyme for catalyzing the conversion of 3-dehydroshikimate (3-DHS) to shikimate (SA); it also potentially participates in GA synthesis in a branch of the SA pathway. In this study, four DQD/SDH proteins were produced in . Three DQD/SDHs had 3-DHS reduction and SA oxidation functions. Notably, three DQD/SDHs showed individual differences between the catalytic efficiency of 3-DHS reduction and SA oxidation; DQD/SDHa had higher catalytic efficiency for 3-DHS reduction than for SA oxidation, DQD/SDHd showed the opposite tendency, and DQD/SDHc had almost equal catalytic efficiency for 3-DHS reduction and SA oxidation. , GA was mainly generated from 3-DHS through nonenzymatic conversion. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) analysis showed that and expression was correlated with GA and 1--galloyl-β-D-glucose accumulation in . These results revealed the DQD/SDHc and DQD/SDHd genes are involved in GA synthesis. Finally, site-directed mutagenesis exhibited the mutation of residues Ser-338 and NRT to Gly and DI/LD in the SDH unit is the reason for the low activity of DQD/SDHb for 3-DHS reduction and SA oxidation.