Gerasymenko, Iryna ; Sheludko, Yuriy V. ; Fuertes, Ismael Navarro ; Schmidts, Volker ; Steinel, Lara ; Haumann, Elisabeth ; Warzecha, Heribert (2022)
Engineering of a plant isoprenyl diphosphate synthase for development of irregular coupling activity.
In: Chembiochem : a European journal of chemical biology, 23 (1)
doi: 10.1002/cbic.202100465
Artikel, Bibliographie

Kurzbeschreibung (Abstract)

We performed mutagenesis on a regular isoprenyl diphosphate synthase (IDS), neryl diphosphate synthase from Solanum lycopersicum (SlNPPS), that has a structurally related analogue performing non-head-to-tail coupling of two dimethylallyl diphosphate (DMAPP) units, lavandulyl diphosphate synthase from Lavandula x intermedia (LiLPPS). Wild-type SlNPPS catalyses regular coupling of isopentenyl diphosphate (IPP) and DMAPP in cis-orientation resulting in the formation of neryl diphosphate. However, if the enzyme is fed with DMAPP only, it is able to catalyse the coupling of two DMAPP units and synthesizes two irregular monoterpene diphosphates; their structures were elucidated by the NMR analysis of their dephosphorylation products. One of the alcohols is lavandulol. The second compound is the trans-isomer of planococcol, the first example of an irregular cyclobutane monoterpene with this stereochemical configuration. The irregular activity of SlNPPS constitutes 0.4% of its regular activity and is revealed only if the enzyme is supplied with DMAPP in absence of IPP. The exchange of asparagine 88 for histidine considerably enhanced the non-head-to-tail coupling. While still only observed in the absence of IPP, irregular activity of the mutant reaches 13.1 % of its regular activity. The obtained results prove that regular IDS are promising starting points for protein engineering aiming at the development of irregular activities and leading to novel monoterpene structures.

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