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J. Biol. Chem., Vol. 276, Issue 25, 22154-22159, June 22, 2001
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From the In higher plants, the de-epoxidation of
violaxanthin (Vx) to antheraxanthin and zeaxanthin is required for the
pH-dependent dissipation of excess light energy as heat and
by that process plays an important role in the protection
against photo-oxidative damage. The de-epoxidation reaction was
investigated in an in vitro system using reconstituted
light-harvesting complex II (LHCII) and a thylakoid raw extract
enriched in the enzyme Vx de-epoxidase. Reconstitution of LHCII with
varying carotenoids was performed to replace lutein and/or neoxanthin,
which are bound to the native complex, by Vx. Recombinant LHCII
containing either 2 lutein and 1 Vx or 1.6 Vx and 1.1 neoxanthin or 2.8 Vx per monomer were studied. Vx de-epoxidation was inducible for all
complexes after the addition of Vx de-epoxidase but to different
extents and with different kinetics in each complex. Analysis of the
kinetics indicated that the three possible Vx binding sites have at
least two, and perhaps three, specific rate constants for
de-epoxidation. In particular, Vx bound to one of the two lutein
binding sites of the native complex, most likely L1, was not at all or
only at a slow rate convertible to Zx. In reisolated LHCII, newly
formed Zx almost stoichiometrically replaced the transformed Vx,
indicating that LHCII and Vx de-epoxidase stayed in close
contact during the de-epoxidation reactions and that no release of
carotenoids occurred.
De-epoxidation of Violaxanthin after Reconstitution into
Different Carotenoid Binding Sites of Light-harvesting Complex II*
§,, Institut für Biochemie der Pflanzen,
Heinrich-Heine-Universität Düsseldorf, D-40225
Düsseldorf, Germany and ¶ Institut für Allgemeine
Botanik, Johannes-Gutenberg-Universität Mainz, Müllerweg 6, D-55128 Mainz, Germany
*
This work was supported by the Deutsche
Forschungsgemeinschaft SFB 189, TP B13, and Ja 665/2-1 (to P. J.)
and Pa-324/5-3 (to H. P.) and a grant from the Stiftung
Rheinland Pfalz für Innovation (to H. P.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
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