Biosynthesis of the Dihydrochalcone Sweetener Trilobatin Requires Phloretin Glycosyltransferase2

• Published in: Plant physiology (Bethesda) 2020-10, Vol.184 (2), p.738-752
• Main Authors: Wang, Yule, Yauk, Yar-Khing, Zhao, Qian, Hamiaux, Cyril, Xiao, Zhengcao, Gunaseelan, Kularajathevan, Zhang, Lei, Tomes, Sumathi, López-Girona, Elena, Cooney, Janine, Li, Houhua, Chagné, David, Ma, Fengwang, Li, Pengmin, Atkinson, Ross G
• Format: Article
• Language: English
• Subjects: Chalcones - metabolism; Flavonoids - biosynthesis; Glycosyltransferases - metabolism; Malus - metabolism; Phloretin - metabolism; Plants, Genetically Modified; Polyphenols - biosynthesis; Sweetening Agents - metabolism
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.20.00807

Epidemics of obesity and type 2 diabetes drive strong consumer interest in plant-based low-calorie sweeteners. Trilobatin is a sweetener found at high concentrations in the leaves of a range of crabapple ( ) species, but not in domesticated apple ( × ) leaves, which contain trilobatin's bitter positional isomer phloridzin. Variation in trilobatin content was mapped to the locus on LG 7 in a segregating population developed from a cross between domesticated apples and crabapples. ( ) was identified by activity-directed protein purification and differential gene expression analysis in samples high in trilobatin but low in phloridzin. Markers developed for cosegregated strictly with the locus. Biochemical analysis showed PGT2 efficiently catalyzed 4'-o-glycosylation of phloretin to trilobatin as well as 3-hydroxyphloretin to sieboldin. Transient expression of double bond reductase, chalcone synthase, and genes reconstituted the apple pathway for trilobatin production in Transgenic × plants overexpressing produced high concentrations of trilobatin in young leaves. Transgenic plants were phenotypically normal, and no differences in disease susceptibility were observed compared to wild-type plants grown under simulated field conditions. Sensory analysis indicated that apple leaf teas from transgenics were readily discriminated from control leaf teas and were perceived as significantly sweeter. Identification of allows marker-aided selection to be developed to breed apples containing trilobatin, and for high amounts of this natural low-calorie sweetener to be produced via biopharming and metabolic engineering in yeast.