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Nucleotide sequence and expression of the mercurial-resistance operon from Staphylococcus aureus plasmid pI258
Abstract:
The mercurial-resistance determinant from Staphylococcus aureus plasmid pI258 is located on a 6.4-kilobase-pair Bgl II fragment. The determinant was cloned into both Bacillus subtilis and Escherichia coli. Mercury resistance was found only in B. subtilis. The 6404-base-pair DNA sequence of the Bgl II fragment was determined. The mer DNA sequence includes seven open reading frames, two of which have been identified by homology with the merA (mercuric reductase) and merB (organomercurial lyase) genes from the mercurial-resistance determinants of Gram-negative bacteria. Whereas 40% of the amino acid residues overall were identical between the pI258 merA polypeptide product and mercuric reductases from Gram-negative bacteria, the percentage identity in the active-site positions and those thought to be involved in NADPH and FAD contacts was above 90%. The 216 amino acid organomercurial lyase sequence was 39% identical with that from a Serratia plasmid, with higher conservation in the middle of the sequences and lower homologies at the amino and carboxyl termini. The remaining five open reading frames in the pI258 mer sequence have no significant homologies with the genes from previously sequenced Gram-negative mer operons.
Insights
The Staphylococcus aureus mercury resistance determinant cloned into Bacillus subtilis showed resistance, unlike in Escherichia coli. DNA sequencing revealed seven open reading frames, including merA and merB homologs, crucial for mercury detoxification.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- The Staphylococcus aureus plasmid pI258 contains a mercury resistance determinant.
- Understanding the genetic basis of mercury resistance is crucial for combating antibiotic resistance.
Purpose of the Study:
- To characterize the mercury resistance determinant from Staphylococcus aureus plasmid pI258.
- To investigate the functional expression of this determinant in different bacterial hosts.
Main Methods:
- Cloning the mercury resistance determinant into Bacillus subtilis and Escherichia coli.
- DNA sequencing of the 6.4-kilobase-pair Bgl II fragment.
- Homology analysis of open reading frames with known mercury resistance genes.
Main Results:
- Mercury resistance was conferred in Bacillus subtilis but not in Escherichia coli.
- DNA sequencing identified seven open reading frames, including homologs of merA (mercuric reductase) and merB (organomercurial lyase).
- High sequence identity (>90%) was observed in active-site regions of the merA homolog compared to Gram-negative bacteria.
Conclusions:
- The Staphylococcus aureus mercury resistance determinant possesses functional merA and merB genes.
- The differential expression or function in B. subtilis and E. coli warrants further investigation.
- Five novel open reading frames suggest unique mechanisms or regulatory elements in this mercury resistance operon.