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Mutsuda, M, Ishikawa, T, Takeda, T and Shigeoka, S (1996) The catalase-peroxidase of Synechococcus PCC 7942: purification, nucleotide sequence analysis and expression in Escherichia coli. Biochem. J. 316 ( Pt 1):251-7

Synechococcus PCC 7942, a cyanobacterium, possesses catalaseperoxidase as the sole hydrogen peroxide-scavenging system. The enzyme has been purified to electrophoretic homogenenity from the cells. The native enzyme had a molecular mass of 150 kDa and was composed of two identical subunits of molecular mass 79 kDa. The apparent Km value of the catalase activity for H2O2 was 4.2 +/- 0.27 mM and the kcat value was 2.6 x 10(4) s-1. The enzyme contained high catalase activity and an appreciable peroxidase activity with o-dianisidine and pyrogallol. The catalase activity was not inhibited by 3-amino-1,2,4-triazole but by KCN and NaN3 (apparent Ki values 19.3 +/- 0.84 and 20.2 +/- 0.95 microM respectively). The enzyme showed an absorption spectrum of typical protohaem and contained one protohaem molecule per dimer. The gene encoding catalase-peroxidase was cloned from the chromosomal DNA of Synechococcus PCC 7942. A 2160 bp open reading frame (ORF), coding a catalase-peroxidase of 720 amino acid residues (approx. 79.9 kDa), was observed. The deduced amino acid sequence coincided with that of the N-terminus of the purified enzyme and showed a remarkable similarity to those of a family of catalase-peroxidases of prokaryotic cells. Escherichia coli BL21 (DE3)plysS, harbouring a recombinant plasmid containing the catalase-peroxidase gene, produced a large amount of proteins that co-migrated on SDS/PAGE with the native enzyme. The recombinant enzyme showed the same ratio of catalase activity to peroxidase activity with o-dianisidine and the same Km for H2O2 as the native enzyme.

PubMed PMC1217331

Amino Acid Sequence; Bacteria/enzymology; Bacterial Proteins; Base Sequence; Chromatography, Gel; Chromatography, Ion Exchange; Cloning, Molecular; Conserved Sequence; Cyanobacteria/enzymology; DNA Primers; Escherichia coli/enzymology; Gene Expression; Kinetics; Macromolecular Substances; Molecular Sequence Data; Molecular Weight; Multienzyme Complexes/chemistry; Multienzyme Complexes/isolation & purification; Multienzyme Complexes/metabolism; Peroxidases/chemistry; Peroxidases/isolation & purification; Peroxidases/metabolism; Polymerase Chain Reaction; Recombinant Proteins/chemistry; Recombinant Proteins/isolation & purification; Recombinant Proteins/metabolism; Sequence Homology, Amino Acid