TY - JOUR
T1 - Molecular basis for branched steviol glucoside biosynthesis
AU - Lee, Soon Goo
AU - Salomon, Eitan
AU - Yu, Oliver
AU - Jez, Joseph M.
N1 - Funding Information:
ACKNOWLEDGMENTS. E.S. was supported by US-Israel Vaadia Binational Agricultural Research Development Fund Postdoctoral Fellowship FI-504-14. Portions of this research were carried out at the Argonne National Laboratory Structural Biology Center of the Advanced Photon Source, a national user facility operated by the University of Chicago by Department of Energy Office of Biological and Environmental Research Grant DE-AC02-06CH11357.
Publisher Copyright:
© 2019 National Academy of Sciences. All rights reserved.
PY - 2019
Y1 - 2019
N2 - Steviol glucosides, such as stevioside and rebaudioside A, are natural products roughly 200-fold sweeter than sugar and are used as natural, noncaloric sweeteners. Biosynthesis of rebaudioside A, and other related stevia glucosides, involves formation of the steviol diterpenoid followed by a series of glycosylations catalyzed by uridine diphosphate (UDP)-dependent glucosyltransferases. UGT76G1 from Stevia rebaudiana catalyzes the formation of the branched-chain glucoside that defines the stevia molecule and is critical for its high-intensity sweetness. Here, we report the 3D structure of the UDP-glucosyltransferase UGT76G1, including a complex of the protein with UDP and rebaudioside A bound in the active site. The X-ray crystal structure and biochemical analysis of site-directed mutants identifies a catalytic histidine and how the acceptor site of UGT76G1 achieves regioselectivity for branched-glucoside synthesis. The active site accommodates a two-glucosyl side chain and provides a site for addition of a third sugar molecule to the C3′ position of the first C13 sugar group of stevioside. This structure provides insight on the glycosylation of other naturally occurring sweeteners, such as the mogrosides from monk fruit, and a possible template for engineering of steviol biosynthesis.
AB - Steviol glucosides, such as stevioside and rebaudioside A, are natural products roughly 200-fold sweeter than sugar and are used as natural, noncaloric sweeteners. Biosynthesis of rebaudioside A, and other related stevia glucosides, involves formation of the steviol diterpenoid followed by a series of glycosylations catalyzed by uridine diphosphate (UDP)-dependent glucosyltransferases. UGT76G1 from Stevia rebaudiana catalyzes the formation of the branched-chain glucoside that defines the stevia molecule and is critical for its high-intensity sweetness. Here, we report the 3D structure of the UDP-glucosyltransferase UGT76G1, including a complex of the protein with UDP and rebaudioside A bound in the active site. The X-ray crystal structure and biochemical analysis of site-directed mutants identifies a catalytic histidine and how the acceptor site of UGT76G1 achieves regioselectivity for branched-glucoside synthesis. The active site accommodates a two-glucosyl side chain and provides a site for addition of a third sugar molecule to the C3′ position of the first C13 sugar group of stevioside. This structure provides insight on the glycosylation of other naturally occurring sweeteners, such as the mogrosides from monk fruit, and a possible template for engineering of steviol biosynthesis.
KW - Lucos ltransferase
KW - Noncaloric sweetener
KW - Plant biochemistr
KW - Stevia
KW - X-ray crystal structure
UR - http://www.scopus.com/inward/record.url?scp=85068192360&partnerID=8YFLogxK
U2 - 10.1073/pnas.1902104116
DO - 10.1073/pnas.1902104116
M3 - Article
C2 - 31182573
AN - SCOPUS:85068192360
SN - 0027-8424
VL - 116
SP - 13131
EP - 13136
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 26
ER -