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Related Experiment Videos

Single-stranded DNA binding proteins (SSBs) from prokaryotic transmissible plasmids.

P P Ruvolo1, K M Keating, K R Williams

  • 1Department of Molecular Biology and Genetics, Albert Einstein College of Medicine, Bronx, New York 10461.

Proteins
|January 1, 1991
PubMed
Summary

Single-stranded DNA binding proteins (SSBs) from conjugative plasmids show conserved DNA-binding residues but diverge in function. F-plasmid SSB is unique, failing to bind DNA or stimulate polymerase activity.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Single-stranded DNA binding proteins (SSBs) are crucial for DNA replication, repair, and recombination.
  • Plasmid-encoded SSBs play vital roles in conjugative transfer and maintaining plasmid stability.
  • Understanding SSB diversity is key to deciphering bacterial DNA metabolism.

Purpose of the Study:

  • To determine and compare the DNA and protein sequences of SSBs from plP71a, plP231a, and R64 conjugative plasmids.
  • To investigate the functional differences and similarities between plasmid SSBs and those from Escherichia coli and F-plasmid.
  • To identify conserved residues involved in DNA binding and explore the role of Tyr-70.

Main Methods:

  • DNA and protein sequence determination and comparative analysis.

Related Experiment Videos

  • In vitro DNA binding studies using poly(dA) and poly[d(A-T)].
  • Enzyme activity assays measuring DNA polymerase III holoenzyme elongation rates.

    • Main Results:

    • High sequence conservation in the N-terminal two-thirds of SSBs, with divergence in the C-terminal third.
    • Conserved DNA-binding residues (Trp-40, Trp-54, Trp-88, His-55, Phe-60) identified across all SSBs.
    • A novel role for Tyr-70 in DNA binding is proposed.
    • F-plasmid SSB demonstrated unique non-binding and non-stimulatory properties.
    • Three plasmid SSBs exhibited superior helix-destabilizing activity compared to E. coli SSB.

    Conclusions:

    • Plasmid SSBs share conserved structural and functional domains, but exhibit functional diversity.
    • The C-terminal region and specific residues are critical for SSB function and DNA interaction.
    • F-plasmid SSB represents a distinct functional variant with implications for plasmid biology.
    • Plasmid SSBs can be more effective helix-destabilizing agents than chromosomal SSBs.