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

被引:185
作者
Kampranis, Sotirios C. [1 ]
Ioannidis, Daphne
Purvis, Alan
Mahrez, Walid
Ninga, Ederina
Katerelos, Nikolaos A.
Anssour, Samir
Dunwell, Jim M.
Degenhardt, Joerg
Makris, Antonios M.
Goodenough, Peter W.
Johnson, Christopher B.
机构
[1] Mediterranean Agron Inst Chania, Dept Nat Prod & Biotechnol, Khania 73100, Greece
[2] Univ Reading, Sch Biol Sci, Reading RG6 6AS, Berks, England
[3] Max Planck Inst Chem Ecol, D-07745 Jena, Germany
关键词
D O I
10.1105/tpc.106.047779
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
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.
引用
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页码:1994 / 2005
页数:12
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