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Molecular cloning and expression of the gene encoding ADP-glucose pyrophosphorylase from the cyanobacteriumAnabaena sp. strain PCC 7120

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Abstract

Previous studies have indicated that ADP-glucose pyrophosphorylase (ADPGlc PPase) from the cyanobacteriumAnabaena sp. strain PCC 7120 is more similar to higher-plant than to enteric bacterial enzymes in antigenicity and allosteric properties. In this paper, we report the isolation of theAnabaena ADPGlc PPase gene and its expression inEscherichia coli. The gene we isolated from a genomic library utilizes GTG as the start codon and codes for a protein of 48347 Da which is in agreement with the molecular mass determined by SDS-PAGE for theAnabaena enzyme. The deduced amino acid sequence is 63, 54, and 33% identical to the rice endosperm small subunit, maize endosperm large subunit, and theE. coli sequences, respectively. Southern analysis indicated that there is only one copy of this gene in theAnabaena genome. The cloned gene encodes an active ADPGlc PPase when expressed in anE. coli mutant strain AC70R1-504 which lacks endogenous activity of the enzyme. The recombinant enzyme is activated and inhibited primarily by 3-phosphoglycerate and Pi, respectively, as is the nativeAnabaena ADPGlc PPase. Immunological and other biochemical studies further confirmed the recombinant enzyme to be theAnabaena enzyme.

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Authors and Affiliations

  1. Dept. of Biochemistry, Michigan State University, 48824, East Lansing, MI, USA

    Yee-yung Charng, Genichi Kakefuda, Alberto A. Iglesias & Jack Preiss

  2. Dept. of Molecular Genetics and Cell Biology, University of Chicago, 60637, IL, USA

    William J. Buikema

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  1. Yee-yung Charng
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  2. Genichi Kakefuda
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  3. Alberto A. Iglesias
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  4. William J. Buikema
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  5. Jack Preiss
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Charng, Yy., Kakefuda, G., Iglesias, A.A. et al. Molecular cloning and expression of the gene encoding ADP-glucose pyrophosphorylase from the cyanobacteriumAnabaena sp. strain PCC 7120. Plant Mol Biol 20, 37–47 (1992). https://doi.org/10.1007/BF00029147

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  • DOI: https://doi.org/10.1007/BF00029147

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