Functional characterizations of ß-glucosidases involved in aroma compound formation in tea (Camellia sinensis).

Tea (Camellia sinensis) aroma is an important factor affecting tea quality. Many tea aroma compounds are present as glycosidically conjugated forms in tea leaves, and can be hydrolyzed by ß-glucosidase (ß-Glu) and ß-primeverosidase to release free tea aromas. ß-Primeverosidase has been identified and functionally characterized, while ß-Glu has not been identified in tea leaves. In the present study, we established a yeast expression system to recombine CsGH1BG1, CsGH3BG1, and CsGH5BG1, which belonged to GH1, GH3, and GH5 families in plants, respectively. These three recombinant Csß-Glus hydrolyzed the ß-glucopyranosidically conjugated aromas to form free aromas, suggesting that there was no specific Csß-Glus for the hydrolysis of ß-glucopyranosidically conjugated aromas in vitro. Furthermore, subcellular localization of the Csß-Glus indicated that CsGH1BG1 and CsGH3BG1 were located in the cytosol and vacuole, respectively, while CsGH5BG1 was located in the cell wall. This suggested that CsGH1BG1 and CsGH3BG1 might be responsible for the hydrolysis of ß-glucopyranosidically conjugated aromas in tea leaves during the tea manufacturing process. This study provides the first evidence of Csß-Glus in tea leaves, and will advance understanding of tea aroma formation.

Formation of (E)-nerolidol in tea (Camellia sinensis) leaves exposed to multiple stresses during tea manufacturing.

(E)-Nerolidol is a volatile sesquiterpene that contributes to the floral aroma of teas (Camellia sinensis). The unique manufacturing process for oolong tea involves multiple stresses, resulting in a high content of (E)-nerolidol, which is not known to form in tea leaves. This study aimed to determine the formation mechanism of (E)-nerolidol in tea exposed to multiple stresses during tea manufacture. C. sinensis (E)-nerolidol synthase (CsNES) recombinant protein, found in the cytosol, was found to transform farnesyl diphosphate into (E)-nerolidol. CsNES was highly expressed during the oolong tea turn over process, resulting in (E)-nerolidol accumulation. Continuous mechanical damage, simulating the turn over process, significantly enhanced CsNES expression level and (E)-nerolidol content. The combination of low temperature stress and mechanical damage had a synergistic effect on (E)-nerolidol formation. This is the first evidence of (E)-nerolidol formation mechanism in tea leaves and a characteristic example of plant volatile formation in response to dual stresses.