Protein domains and conformational changes in the activation of RepA, a DNA replication initiator.
RepA is the DNA replication initiator protein of the Pseudomonas plasmid pPS10. RepA has a dual function: as a dimer, it binds to an inversely-repeated sequence acting as a repressor of its own synthesis; as a monomer, RepA binds to four directly-repeated sequences to constitute a specialized nucleoprotein complex responsible for the initiation of DNA replication. We have previously shown that a Leucine Zipper-like motif (LZ) at the N-terminus of RepA is responsible for protein dimerization. In this paper we characterize the existence in RepA of two protein globular domains C-terminal to the LZ. We propose that dissociation of RepA dimers into monomers results in a conformational change from a compact arrangement of both domains, competent for binding to the operator, to an extended species that is suited for iteron binding. This model establishes the structural basis for the activation of DNA replication initiators in plasmids from Gram-negative bacteria.
Departmento de Microbiología Molecular, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas (CSIC), Madrid, Spain.
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Giraldo
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Andreu
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Díaz-Orejas
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002850:Chromatography, Gel; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018983:DNA Footprinting; D004265:DNA Helicases; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D017665:Hydroxyl Radical; D009875:Operator Regions, Genetic; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014176:Protein Biosynthesis; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000235:genetics; D011549:Pseudomonas; D012093:Replicon; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D015534:Trans-Activators; D014364:Tryptophan / Q000737:chemistry
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D010446:Peptide Fragments; D011506:Proteins; D015534:Trans-Activators; C045264:replication initiator protein; D017665:Hydroxyl Radical; D014364:Tryptophan; D004265:DNA Helicases