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Abstract:
The cloned genetic determinant of R16 (IncB) plasmid primase encodes two polypeptides (apparent Mr 240,000 and 175,000), probably products of the same coding sequence of DNA, which are also detectable in extracts of cells carrying the parent plasmid. Deletion analysis and transposon mutagenesis indicate that only about 1.7 kilobase pairs (kb) of the cloned fragment, encoding a truncated polypeptide of 76,000 Da, is necessary for activity. The cloned genetic determinant of the R387 (IncK) plasmid primase, which encodes polypeptides of apparent Mr 270,000 and 200,000, appears to be very similar to the R16 gene except for an additional sequence of approximately 0.65 kb.
Insights
The R16 (IncB) and R387 (IncK) plasmid primase genes were analyzed. A minimal 1.7 kb fragment of the R16 primase gene is sufficient for activity, showing similarity to the R387 gene.
Area of Science:
- Molecular Biology
- Genetics
- Microbiology
Background:
- Plasmid replication relies on primase enzymes.
- The R16 (IncB) and R387 (IncK) plasmids are important in bacterial genetics.
Purpose of the Study:
- To characterize the genetic determinants of R16 and R387 plasmid primases.
- To identify the minimal DNA sequence required for primase activity.
Main Methods:
- Cloning of plasmid DNA fragments.
- Deletion analysis to determine essential regions.
- Transposon mutagenesis to identify functional domains.
Main Results:
- The R16 primase determinant encodes polypeptides of 240,000 and 175,000 Da.
- A 1.7 kb fragment of the R16 determinant, encoding a 76,000 Da polypeptide, is sufficient for primase activity.
- The R387 primase determinant is similar to R16 but contains an additional 0.65 kb sequence.
Conclusions:
- The minimal functional unit of the R16 plasmid primase has been identified.
- Structural similarities and differences between R16 and R387 primase determinants were elucidated.