摘要
Diterpene glycosyltransferase UGT76G1 from Stevia rebaudiana (SrUGT76G1) is key to the generation ofeconomically important steviol glycosides (SGs), a group of natural sweeteners with high-intensity sweetness. SrUGT76G1 accommodates a wide range of steviol-derived substrates and many other small molecules. We report here the crystal structures of SrUGT76G1 in complex with multiple ligands to answer howthis enzyme recognizes diterpenoid aglycones and catalyzes the 1,3-sugar chain branching. A spaciouspocket for sugar-acceptor binding was observed from the determined SrUGT76G1 structures, which canexplain its broad substrate spectrum. Residues Gly87 and Leu204 lining the pocket play key roles in switching between diterpenoid and flavonoid glucosylation. An engineered mutant of SrUGT76G1, T284S, couldcatalyze a selectively increased production of next-generation sweetener rebaudioside M, with diminishedside product of rebaudioside I. Taken together, these resutls provide significant insights into molecularbasis of the substrate specificity of scarcely documented diterpenoid glycosyltransferases and wouldfacilitate the structure-guided glycoengineering to produce diversified diterpenoids with new activities.
基金
This work was financially supported by the National Key R&D Program of China(2018YFA0900600)
the Strategic Priority Research Program "Molecular Mechanism of Plant Growth and Development" of CAS(XDB27020202,XDB27020103)
the National Natural Science Foundation of China(31700263,31670099,31700261)
grants from the Shanghai Science and Technology Commission(19XD1424500)
J.L.is supported by the Foundation of Youth Innovation Promotion Association of the Chinese Academy of Sciences
This work was also financially supported by the Construction of the Registry and Database of Bioparts for Synthetic Biology of the Chinese Academy of Science(no.ZSYS-016)
the International Partnership Program of Chinese Academy of Science(no.153D31KYSB20170121)
the National Key Laboratory of Plant Molecular Genetics,SIPPI,CAS.
