Intramolecular proton transfer in the cyclization of geranylgeranyl diphosphate to the taxadiene precursor of taxol catalyzed by recombinant taxadiene synthase.
BACKGROUND
The committed step in the biosynthesis of the anticancer drug taxol in yew (Taxus) species is the cyclization of geranylgeranyl diphosphate to taxa-4(5),11(12)-diene. The enzyme taxadiene synthase catalyzes this complex olefin cation cyclization cascade involving the formation of three rings and three stereogenic centers.


RESULTS
Recombinant taxadiene synthase was incubated with specifically deuterated substrates, and the mechanism of cyclization was probed using MS and NMR analyses of the products to define the crucial hydrogen migration and terminating deprotonation steps. The electrophilic cyclization involves the ionization of the diphosphate with closure of the A-ring, followed by a unique intramolecular transfer of the C11 proton to the re-face of C7 to promote closure of the B/C-ring juncture, and cascade termination by proton elimination from the beta-face of C5.


CONCLUSIONS
These findings provide insight into the molecular architecture of the first dedicated step of taxol biosynthesis that creates the taxane carbon skeleton, and they have broad implications for the general mechanistic capability of the large family of terpenoid cyclization enzymes.
Williams
D C
DC
Institute of Biological Chemistry, Washington State University, Pullman, WA 99164-6340, USA.
Carroll
B J
BJ
Jin
Q
Q
Rithner
C D
CD
Lenger
S R
SR
Floss
H G
HG
Coates
R M
RM
Williams
R M
RM
Croteau
R
R
D000475:Alkenes / Q000737:chemistry / Q000378:metabolism*; D000972:Antineoplastic Agents, Phytogenic / Q000378:metabolism; D002412:Cations; D003500:Cyclization; D003903:Deuterium; D004224:Diterpenes / Q000737:chemistry / Q000378:metabolism*; D007535:Isomerases / Q000235:genetics / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008968:Molecular Conformation; D015394:Molecular Structure; D017239:Paclitaxel / Q000096:biosynthesis* / Q000737:chemistry; D010946:Plants, Medicinal; D011106:Polyisoprenyl Phosphates / Q000737:chemistry / Q000378:metabolism*; D011522:Protons; D011994:Recombinant Proteins / Q000378:metabolism; D020946:Taxus / Q000201:enzymology* / Q000235:genetics
D000475:Alkenes; D000972:Antineoplastic Agents, Phytogenic; D002412:Cations; D004224:Diterpenes; D011106:Polyisoprenyl Phosphates; D011522:Protons; D011994:Recombinant Proteins; C093125:taxa-4(5),11(12)diene; D003903:Deuterium; D007535:Isomerases; C095926:taxa-4(5),11(12)-diene synthase; C002963:geranylgeranyl pyrophosphate; D017239:Paclitaxel