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Existence of two ferredoxin-glutamate synthases in the cyanobacterium Synechocystis sp. PCC 6803. Isolation and insertional inactivation of gltB and gltS genes

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Abstract

The first two genes of ferredoxin-dependent glutamate synthase (Fd-GOGAT) from a prokaryotic organism, the cyanobacterium Synechocystis sp. PCC 6803, were cloned in Escherichia coli. Partial sequencing of the cloned genomic DNA, of the 6.3 kb Hind III and 9.3 kb Cla I fragments, confirmed the existence of two different genes coding for glutamate synthases, named gltB and gltS. The gltB gene was completely sequenced and encodes for a polypeptide of 1550 amino acid residues (M r 168 964). Comparative analysis of the gltB deduced amino acid sequence against other glutamate synthases shows a higher identity with the alfalfa NADH-GOGAT (55.2%) than with the corresponding Fd-GOGAT from the higher plants maize and spinach (about 43%), the red alga Antithamnnion sp. (42%) or with the NADPH-GOGAT of bacterial source, such as Escherichia coli (41%) and Azospirillum brasilense (45%). The detailed analysis of Synechocystis gltB deduced amino acid sequence shows strongly conserved regions that have been assigned to the 3Fe-4S cluster (CX5CHX3C), the FMN-binding domain and the glutamine-amide transferase domain. Insertional inactivation of gltB and gltS genes revealed that both genes code for ferredoxin-dependent glutamate synthases which were nonessential for Synechocystis growth, as shown by the ferredoxin-dependent glutamate synthase activity and western-blot analysis of the mutant strains.

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Authors and Affiliations

  1. Instituto de Bioquímica Vegetal y Fotosíntesis, Universidad de Sevilla-CSIC, Apdo. 1113, 41080, Sevilla, Spain

    Francisco Navarro, Sebastián Chávez, Pedro Candau & Francisco J. Florencio

  2. Departamento de Bioquímica Vegetal y Biología Molecular, Universidad de Sevilla-CSIC, Apdo. 1113, 41080, Sevilla, Spain

    Francisco Navarro, Sebastián Chávez, Pedro Candau & Francisco J. Florencio

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  1. Francisco Navarro
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  4. Francisco J. Florencio
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Navarro, F., Chávez, S., Candau, P. et al. Existence of two ferredoxin-glutamate synthases in the cyanobacterium Synechocystis sp. PCC 6803. Isolation and insertional inactivation of gltB and gltS genes. Plant Mol Biol 27, 753–767 (1995). https://doi.org/10.1007/BF00020228

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