Induced Bending of Plasmid pLS1 DNA by the Plasmid-encoded Protein RepA

The broad host range streptococcal plasmid pLS1 encodes for a 5.1-kDa repressor protein, RepA. This protein has affinity for DNA (linear or supercoiled) and is translated from the same mRNA as the replication initiator protein RepB. By gel retardation assays, we observed that RepA shows specificity...

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Bibliographic Details
Published in:The Journal of biological chemistry 1989-12, Vol.264 (35), p.21334-21339
Main Authors: Pérez-Martín, J, del Solar, G H, Lurz, R, de la Campa, A G, Dobrinski, B, Espinosa, M
Format: Article
Language:English
Subjects:
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Base Sequence
Biological and medical sciences
Cloning, Molecular - methods
Computer Simulation
DNA Helicases
DNA Replication
DNA, Bacterial - genetics
DNA, Bacterial - metabolism
DNA, Bacterial - ultrastructure
DNA-Binding Proteins
Fundamental and applied biological sciences. Psychology
Interactions. Associations
Intermolecular phenomena
Microscopy, Electron
Molecular biophysics
Molecular Sequence Data
Nucleic Acid Conformation
Plasmids
Proteins
Restriction Mapping
Streptococcus pneumoniae - genetics
Substrate Specificity
Trans-Activators
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Summary:The broad host range streptococcal plasmid pLS1 encodes for a 5.1-kDa repressor protein, RepA. This protein has affinity for DNA (linear or supercoiled) and is translated from the same mRNA as the replication initiator protein RepB. By gel retardation assays, we observed that RepA shows specificity for binding to the plasmid HinfID fragment, which includes the target of the protein. The target of RepA within the plasmid DNA molecule has been located around the plasmid single site ApaLI. This site is included in a region that contains the promoter for the repA and repB genes and is contiguous to the plasmid ori(+). A complex sequence-directed DNA curvature is observed in this region of pLS1. Upon addition of RepA to plasmid linear DNA or to circularly permuted restriction fragments, this intrinsic curvature was greatly enhanced. Thus, a strong RepA-induced bending could be located in the vicinity of the ApaLI site. Visualization of the bent DNA was achieved by electron microscopy of complexes between RepA and plasmid DNA fragments containing the RepA target.
ISSN:0021-9258
1083-351X
DOI:10.1016/S0021-9258(19)30084-5