TY - JOUR
T1 - Rational conversion of substrate and product specificity in a Salvia monoterpene synthase
T2 - structural insights into the evolution of terpene synthase function
AU - Kampranis, Sotirios
AU - Ioannidis, Daphne
AU - Purvis, Alan
AU - Mahrez, Walid
AU - Ninga, Ederina
AU - Katerelos, Nikolaos A
AU - Anssour, Samir
AU - Dunwell, Jim M
AU - Degenhardt, Jörg
AU - Makris, Antonios M
AU - Goodenough, Peter W
AU - Johnson, Christopher B
PY - 2007/6
Y1 - 2007/6
N2 - Terpene synthases are responsible for the biosynthesis of the complex chemical defense arsenal of plants and microorganisms. How do these enzymes, which all appear to share a common terpene synthase fold, specify the many different products made almost entirely from one of only three substrates? Elucidation of the structure of 1,8-cineole synthase from Salvia fruticosa (Sf-CinS1) combined with analysis of functional and phylogenetic relationships of enzymes within Salvia species identified active-site residues responsible for product specificity. Thus, Sf-CinS1 was successfully converted to a sabinene synthase with a minimum number of rationally predicted substitutions, while identification of the Asn side chain essential for water activation introduced 1,8-cineole and alpha-terpineol activity to Salvia pomifera sabinene synthase. A major contribution to product specificity in Sf-CinS1 appears to come from a local deformation within one of the helices forming the active site. This deformation is observed in all other mono- or sesquiterpene structures available, pointing to a conserved mechanism. Moreover, a single amino acid substitution enlarged the active-site cavity enough to accommodate the larger farnesyl pyrophosphate substrate and led to the efficient synthesis of sesquiterpenes, while alternate single substitutions of this critical amino acid yielded five additional terpene synthases.
AB - Terpene synthases are responsible for the biosynthesis of the complex chemical defense arsenal of plants and microorganisms. How do these enzymes, which all appear to share a common terpene synthase fold, specify the many different products made almost entirely from one of only three substrates? Elucidation of the structure of 1,8-cineole synthase from Salvia fruticosa (Sf-CinS1) combined with analysis of functional and phylogenetic relationships of enzymes within Salvia species identified active-site residues responsible for product specificity. Thus, Sf-CinS1 was successfully converted to a sabinene synthase with a minimum number of rationally predicted substitutions, while identification of the Asn side chain essential for water activation introduced 1,8-cineole and alpha-terpineol activity to Salvia pomifera sabinene synthase. A major contribution to product specificity in Sf-CinS1 appears to come from a local deformation within one of the helices forming the active site. This deformation is observed in all other mono- or sesquiterpene structures available, pointing to a conserved mechanism. Moreover, a single amino acid substitution enlarged the active-site cavity enough to accommodate the larger farnesyl pyrophosphate substrate and led to the efficient synthesis of sesquiterpenes, while alternate single substitutions of this critical amino acid yielded five additional terpene synthases.
KW - Alkyl and Aryl Transferases
KW - Amino Acid Sequence
KW - Binding Sites
KW - Chromatography, Gas
KW - Crystallography, X-Ray
KW - Evolution, Molecular
KW - Molecular Sequence Data
KW - Mutation
KW - Protein Structure, Secondary
KW - Salvia
KW - Structure-Activity Relationship
KW - Substrate Specificity
KW - Volatilization
U2 - 10.1105/tpc.106.047779
DO - 10.1105/tpc.106.047779
M3 - Journal article
C2 - 17557809
SN - 1040-4651
VL - 19
SP - 1994
EP - 2005
JO - Plant Cell
JF - Plant Cell
IS - 6
ER -