Plant Physiology Preview
Published on November 19, 2004; 10.1104/pp.104.052092
Received August 19, 2004
Returned for revision October 16, 2004
Accepted October 17, 2004
Analysis in Vitro of the Enzyme CRTISO Establishes a Poly-cis-Carotenoid Biosynthesis Pathway in Plants
Tal Isaacson , Itzhak Ohad , Peter Beyer , and Joseph Hirschberg *
Departments of Genetics, The Hebrew University of Jerusalem, Jerusalem 91904, Israel
Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem 91904, Israel
Center for Applied Biosciences, Universität Freiburg, 79104 Freiburg, Germany
* Corresponding author; email: hirschu{at}vms.huji.ac.il.
Most enzymes in the central pathway of carotenoid biosynthesis in plants have been identified and studied at the molecular level. However, the specificity and role of cis-trans-isomerization of carotenoids, which occurs in vivo during carotene biosynthesis, remained unresolved. We have previously cloned from tomato (Solanum lycopersicum) the CrtISO gene, which encodes a carotene cis-trans-isomerase. To study the biochemical properties of the enzyme, we developed an enzymatic in vitro assay in which a purified tomato CRTISO polypeptide overexpressed in Escherichia coli cells is active in the presence of an E. coli lysate that includes membranes. We show that CRTISO is an authentic carotene isomerase. Its catalytic activity of cis-to-trans isomerization requires redox-active components, suggesting that isomerization is achieved by a reversible redox reaction acting at specific double bonds. Our data demonstrate that CRTISO isomerizes adjacent cis-double bonds at C7 and C9 pairwise into the trans-configuration, but is incapable of isomerizing single cis-double bonds at C9 and C9'. We conclude that CRTISO functions in the carotenoid biosynthesis pathway in parallel with  -carotene desaturation, by converting 7,9,9'-tri-cis-neurosporene to 9'-cis-neurosporene and 7'9'-di-cis-lycopene into all-trans-lycopene. These results establish that in plants carotene desaturation to lycopene proceeds via cis-carotene intermediates.
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