Functional analyses of Tyr420 and Leu607 of Alicyclobacillus acidocaldarius squalene-hopene cyclase. Neoachillapentaene, a novel triterpene with the 1, 5, 6-trimethylcyclohexene moiety produced through a folding of the constrained boat structure.
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The functions of Tyr420 and Leu607 were analyzed by constructing various site-directed mutants. The mutation at position 420 into Ala and Gly gave bicyclic α- and γ-polypodatetraene in significant amounts, but with a trace amount of tricyclic malabaricatriene. The kinetic data for and the product distribution of the Y420F mutant indicate that the major function of Tyr420 is to stabilize the 6/6-fused bicyclic cation. Mutation experiments on Leu607 demonstrate that the appropriate steric bulk size at position 607 is required to strongly bind with the product-like conformation formed during the polycyclization process. Introduction of the bulkiest Tip residue into 420 or 607 led to the production of a novel monocyclic triterpene having the (5R, 6R)-1,5,6-trimethylcyclohexene ring, named neoachillapentaene,indicating that the enzymatic cyclization proceeded via a constrained boat structure. Folding of the squalene molecule into a boat conformation by squalene cyclase has not been reported before.
雑誌名
Bioscience, Biotechnology, and Biochemistry
巻
66
号
8
ページ
1660 - 1670
発行年
2002
出版者
Japan Society for Bioscience, Biotechnology, and Agrochemistry
Functional analyses of Tyr420 and Leu607 of Alicyclobacillus acidocaldarius squalene-hopene cyclase. Neoachillapentaene, a novel triterpene with the 1, 5, 6-trimethylcyclohexene moiety produced through a folding of the constrained boat structure.
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