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Issue 23, 2014
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Improved selectivity of an engineered multi-product terpene synthase

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Abstract

Mutation of the sesquiterpene synthase Cop2 was conducted with a high-throughput screen for the cyclization activity using a non-natural substrate. A mutant of Cop2 was identified that contained three amino acid substitutions. This mutant, 17H2, converted the natural substrate FPP into germacrene D-4-ol with 77% selectivity. This selectivity is in contrast to that of the parent enzyme in which germacrene D-4-ol is produced as 29% and α-cadinol is produced as 46% of the product mixture. The mutations were shown to each contribute to this selectivity, and a homology model suggested that the mutations lie near to the active site though would be unlikely to be targeted for mutation by rational methods. Kinetic comparisons show that 17H2 maintains a kcat/KM of 0.62 mM−1 s−1, which is nearly identical to that of the parent Cop2, which had a kcat/KM of 0.58 mM−1 s−1.

Graphical abstract: Improved selectivity of an engineered multi-product terpene synthase

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Publication details

The article was received on 03 Mar 2014, accepted on 09 Apr 2014 and first published on 10 Apr 2014


Article type: Paper
DOI: 10.1039/C4OB00479E
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Citation: Org. Biomol. Chem., 2014,12, 4013-4020
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    Improved selectivity of an engineered multi-product terpene synthase

    R. Lauchli, J. Pitzer, R. Z. Kitto, K. Z. Kalbarczyk and K. S. Rabe, Org. Biomol. Chem., 2014, 12, 4013
    DOI: 10.1039/C4OB00479E

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