The cis-iridodial and trans-iridodial freely interconverted with
cis-trans-nepetalactol12, and although CYP76A26 seemed to use
the bicyclic nepetalactol as the preferred substrate, the
monocyclic cis- and trans-iridodials were also utilized, possibly
after spontaneous conversion into nepetalactol (Fig. 4). The
interconversion and sequential metabolism of nepetalactol and
the iridodials in aqueous solution prevented reliable evaluation
of the catalytic parameters with these substrates. Iridotrial
was previously proposed as an intermediate of the secologanin
pathway8. Whereas we never detected iridotrial as an
intermediate in the iridodial to 7-deoxyloganetic acid
conversion under conditions more likely to capture early
reaction products such as low substrate concentrations, short
incubations or low temperature, the latter was very efficiently
converted into 7-deoxyloganetic acid with a Kmapp of 25 mM
and kcat of 5.2 s1 (Supplementary Fig. 3). This suggests that
CYP76A26 is a multifunctional P450 enzyme catalysing
successive hydroxylation/dehydration steps via a mechanism
similar to that recently described for CYP701A3, which catalyses
the conversion of kaurene to kaurenoic acid in the biosynthesis of
gibberellins22
The cis-iridodial and trans-iridodial freely interconverted withcis-trans-nepetalactol12, and although CYP76A26 seemed to usethe bicyclic nepetalactol as the preferred substrate, themonocyclic cis- and trans-iridodials were also utilized, possiblyafter spontaneous conversion into nepetalactol (Fig. 4). Theinterconversion and sequential metabolism of nepetalactol andthe iridodials in aqueous solution prevented reliable evaluationof the catalytic parameters with these substrates. Iridotrialwas previously proposed as an intermediate of the secologaninpathway8. Whereas we never detected iridotrial as anintermediate in the iridodial to 7-deoxyloganetic acidconversion under conditions more likely to capture earlyreaction products such as low substrate concentrations, shortincubations or low temperature, the latter was very efficientlyconverted into 7-deoxyloganetic acid with a Kmapp of 25 mMand kcat of 5.2 s1 (Supplementary Fig. 3). This suggests thatCYP76A26 is a multifunctional P450 enzyme catalysingsuccessive hydroxylation/dehydration steps via a mechanismsimilar to that recently described for CYP701A3, which catalysesthe conversion of kaurene to kaurenoic acid in the biosynthesis ofgibberellins22
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