Rational conversion of substrate and product specificity in a Salvia monoterpene synthase: structural insights into the evolution of terpene synthase function

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 -