Cloning and Characterisation of Chlorophyll Synthase from Avena sativa
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Heidi C. Schmid
Abstract
The chlorophyll synthase gene from oat (Avena sativa) was cloned and expressed in Escherichia coli. The deduced amino acid sequence consists of 378 amino acids including a presequence of 46 amino acids. Deletion mutants show that a core protein comprising amino acid residues 88 to 377 is enzymatically active. The sequence of the mature protein shows 85% identity with the chlorophyll synthase of Arabidopsis thaliana and 62% identity with the chlorophyll synthase of Synechocystis PCC 6803. The gene is constitutively expressed as the same transcript level is found in darkgrown and in lightgrown seedlings. The enzyme requires magnesium ions for activity; manganese ions can reconstitute only part of the activity. Diacetyl and Nphenylmaleimide (NPM) inhibit the enzyme activity. Sitedirected mutagenesis reveals that, out of the 4 Arg residues present in the active core protein, Arg-91 and Arg-161 are essential for the activity. Five cysteine residues are present in the core protein, of which only Cys-109 is essential for the enzyme activity. Since the wildtype and all other Cysmutants with the exception of the mutant C304A are inhibited by Nphenylmaleimide, we conclude that the inhibitor binds to a nonessential Cys residue to abolish activity. The role of the various Arg and Cys residues is discussed.
Copyright © 2001 by Walter de Gruyter GmbH & Co. KG
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Articles in the same Issue
- Biological Activity of Mammalian Transcriptional Repressors
- Cloning and Characterisation of Chlorophyll Synthase from Avena sativa
- DNA Damage by 3,6-Dihydropyrazine-2,5-Dipropanoic Acid, the Cyclic Dimerization Product of 5-Aminolevulinic Acid
- Chicken Erythrocyte Pyrimidine 5-Nucleotidase: Purification and Characterization of the Subclass I Enzyme
- Coenzymes Q9 and Q10, Vitamin E and Peroxidation in Rat Synaptic and Non-Synaptic Occipital Cerebral Cortex Mitochondria during Ageing
- Membrane Activity of (Cys48Ser) Lung Surfactant Protein B Increases with Dimerisation
- Participation of Residue F552 in Domain III of the Protective Antigen in the Biological Activity of Anthrax Lethal Toxin
- Purification of Chrysancorin, a Novel Antifungal Protein with Mitogenic Activity from Garland Chrysanthemum Seeds
- Legumain Forms from Plants and Animals Differ in Their Specificity
- Assignment of the Complete Disulphide Bridge Pattern in the Human Recombinant Follitropin β -Chain
- The Small GTPase Ras Is Involved in Growth Factor-Regulated Expression of the α1 Integrin Subunit in PC12 Cells
- The Mouse Gap Junction Gene Connexin29 Is Highly Expressed in Sciatic Nerve and Regulated during Brain Development
- A Selenocysteine-Containing Peroxiredoxin from the Strictly Anaerobic Organism Eubacterium acidaminophilum
- Characterization of C-Terminally Truncated Human Tissue Inhibitor of Metalloproteinases-4 Expressed in Pichia pastoris
