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|Title:||Localization, biochemical characterization, and solubilization of (4-vinyl) chlorophyllide a reductase, a novel chlorophyll a biosynthetic enzyme|
|Doctoral Committee Chair(s):||Rebeiz, Constantin A.|
|Department / Program:||Plant Biology|
|Degree Granting Institution:||University of Illinois at Urbana-Champaign|
|Abstract:||(4-vinyl) chlorophyllide a reductase (4VCR), catalyses the conversion of divinyl chlorophyllide a (DVChlide a) to monovinyl chlorophyllide a (MVChlide a). The latter is the immediate precursor of monovinyl chlorophyll a (MVChl a) in plants and algae. In reaction center and light harvesting pigment-protein complexes, MVChl a is the main, photosynthetically active protein.
4VCR has been localized in membrane fractions of plastids isolated from etiolated cucumber (Cucumis sativus L.) cotyledons. The enzyme has also been detected in two monocotyledonous species, namely corn (Zea mays L.), and barley (Hordeum vulgare L.). In these species, 4VCR activity has been shown to be maximal in etiolated tissues and drop sharply upon illumination. 4VCR requires the reducing power NADPH as source or hydride ion. Neither a metal ion, nor any cofactor requirement, other than NADPH, could be demonstrated for 4VCR activity. The latter has been shown to be optimal at 30$\sp\circ$C, pH 6.3. Michaelis-Menten constants for DVChlide a and NADPH have been determined to be respectively 0.09-2.00 $\mu$M, and 1.1 mM. 4VCR's low Km value for DVChlide a, in parallel with substrate specificity studies suggested that the enzyme is highly specific for DVChlide a and not other, early and late, divinyl intermediates. The narrow substrate specificity of this bridge-enzyme contrasts with the broad specificity of the forward enzymes of the pathway. The diphenyl ether herbicide acifluorfen methyl (AFM), and the photodynamic herbicide modulator 1, 10-phenanthroline (OPh), inhibited 4VCR activity with respective Ki values of 0.86 and 1.47 mM. The structural requirements of this inhibition have been demonstrated to lie on the presence of at least one nitrogen in a 3-ring, heterocyclic, phenanthroline-related inhibitor. Moreover, electrostatic interactions involving the fractional charge at position 7 of the inhibitor heterocycle appear to be at play in the binding of the inhibitor to the enzyme. Preliminary structural studies of the enzyme have revealed the presence of free, and accessible cystein residues, essential for enzyme activity. Finally, strong inhibition caused by the flavine antagonist quinacrine, suggested that a flavine nucleotide may be involved in the transfer of the hydride from NADPH to DVChlide a.
|Rights Information:||Copyright 1994 Parham, Ramin|
|Date Available in IDEALS:||2011-05-07|
|Identifier in Online Catalog:||AAI9512507|
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