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Identification and Characterization of the sodA Genes Encoding Manganese Superoxide Dismutases in Vibrio parahaemolyticus, Vibrio mimicus , and Vibrio vulnificus
Author(s) -
Kimoto Ryoko,
Funahashi Tatsuya,
Yamamoto Noriko,
Miyoshi Shinichi,
Narimatsu Shizuo,
Yamamoto Shigeo
Publication year - 2001
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.2001.tb01281.x
Subject(s) - vibrio parahaemolyticus , biology , vibrio vulnificus , open reading frame , escherichia coli , gene , peptide sequence , vibrio , nucleic acid sequence , microbiology and biotechnology , biochemistry , operon , gel electrophoresis , amino acid , sequence analysis , bacteria , genetics
Sequencing of Fur titration assay‐positive clones obtained from genomic DNA libraries of Vibrio parahaemolyticus, V. mimicus and V. vulnificus revealed open reading frames encoding proteins of 202, 205 and 202 amino acid residues, respectively. Each open reading frame was preceded by a predicted Fur box which overlaps a likely promoter with similarity to the −10 and −35 consensus sequence of Escherichia coli . The deduced amino acid sequences shared considerable homology with bacterial Mn‐containing superoxide dismutases (MnSODs). Consistent with this, these Vibrio strains produced proteins with SOD activity resistant to inhibition by H 2 O 2 and KCN only when grown under iron‐limiting conditions. Primer extension analysis of the total RNA from these vibrios revealed iron‐repressible expression of the genes. Furthermore, when grown under iron‐limiting conditions, E. coli carrying a plasmid with each cloned gene over‐expressed protein with the same electrophoretic mobility and insensitivity of SOD activity to H 2 O 2 and KCN. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis followed by N‐terminal amino acid sequencing revealed that proteins (MnSODs) having N‐terminal amino acid sequences consistent with those deduced from the corresponding genes were present in cell lysates of the vibrios grown under these iron‐limited conditions. These results demonstrate that the genes cloned in this study are sodA homologs encoding MnSODs, whose expression is regulated by the iron status of the growth medium. PCR using a primer set based on the V. parahaemolyticus sodA sequence revealed the presence of homologous genes in certain other Vibrio species.