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DNA sequence and expression of a defective mer operon from Pseudomonas K-62 plasmid pMR26
1Faculty of Pharmaceutical Sciences, Setsunan University, Hirakata, Osaka, Japan.
Abstract:
pMRB01 cloned from Pseudomonas K-62 plasmid pMR26 conferred bacterial hypersensitivity to organomercurials. DNA sequence analysis of a 2.3-kb SacI-Aor51HI fragment encompassing the whole region required for expression of the hypersensitive phenotype, revealed three open reading frames. The DNA sequence of these frames had 82.5%, 99.2% and 97.0% homology with the pDU1358 merR, merB and merD, respectively. The pMRB01 mer operon differs from the already known mer operon by the absence of the merT, merP and merA genes in this plasmid. An inverted repeat-like sequence upstream from the predicted merR was observed suggesting that this defective mer operon could be part of a transposon-like structure. Induction experiments and maxicell analysis of the mer-polypeptide showed that the lyase enzyme encoded by pMRB01 merB gene is mercurial-inducible and regulated by the transacting product of the merR gene. These results suggest that the hypersensitivity to organomercurials resulted from the expression of lyase activity encoded by the defective mer operon in the absence of reductase activity. The lyase enzyme encoded by pMRB01 merB catalyzes the protonolysis of the C-Hg bond of both arylmercury and alkylmercury compounds.
Insights
This study reveals a defective mercury operon in Pseudomonas K-62, conferring hypersensitivity to organomercurials. The operon expresses a mercurial-inducible lyase enzyme, leading to increased bacterial sensitivity.
Area of Science:
- Microbiology
- Molecular Biology
- Environmental Science
Background:
- Organomercurial compounds are environmental pollutants with significant toxicity.
- Bacterial resistance mechanisms often involve mercury detoxification pathways.
- Understanding novel mercury resistance genes is crucial for bioremediation strategies.
Purpose of the Study:
- To characterize a novel mercury resistance determinant, pMRB01, from Pseudomonas K-62.
- To elucidate the genetic organization and functional properties of the pMRB01 mercury operon.
- To investigate the mechanism underlying bacterial hypersensitivity to organomercurials conferred by pMRB01.
Main Methods:
- DNA sequencing and analysis of a 2.3-kb fragment from plasmid pMRB01.
- Homology comparisons with known mercury resistance genes (mer operon).
- Induction experiments and maxicell analysis to determine gene expression and protein function.
Main Results:
- A defective mercury operon (merR, merB, merD) was identified in pMRB01, lacking merT, merP, and merA genes.
- The pMRB01 merB gene encodes a mercurial-inducible lyase enzyme regulated by the merR gene product.
- Bacterial hypersensitivity to organomercurials is attributed to lyase activity in the absence of reductase activity.
Conclusions:
- The pMRB01 mercury operon represents a novel, defective system conferring organomercurial hypersensitivity.
- The lyase enzyme encoded by pMRB01 merB actively cleaves C-Hg bonds in organomercurials.
- This defective operon, potentially part of a transposon-like structure, offers insights into mercury metabolism and bacterial adaptation.