Cloning, characterization, and nucleotide sequence analysis of the argH gene from Campylobacter jejuni TGH9011 encoding argininosuccinate lyase.

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RESUMO

The complete structural gene for argininosuccinate lyase (argH) from Campylobacter jejuni TGH9011 has been cloned into Escherichia coli by complementation of an E. coli argH auxotrophic mutant. The gene has been subcloned for sequencing on a 4.1-kb DNA segment and localized by the complementing activity of deletion mutants. The complete DNA sequence of the C. jejuni argH gene was determined. The transcription start point for argH mRNA was determined by primer extension analysis and found to be within the coding sequence of the upstream gene, identified as the phosphoenolpyruvate carboxykinase gene (ppc). The argininosuccinate lyase and the phosphoenolpyruvate carboxykinase reading frames overlap by one base, the second example of this phenomenon in C. jejuni chromosomal genes. The enzyme has a deduced subunit molecular weight of 51,831. Recombinant plasmids containing the argH gene generate a 56-kDa protein and a 43-kDa protein in E. coli maxicells. An alternate translation initiation producing a polypeptide with a deduced molecular mass of 42 kDa may account for the smaller protein observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The C. jejuni argH gene shows nucleotide homology to both yeast and human argininosuccinate lyase genes, and conserved amino acid domains are evident between the corresponding proteins.

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