Uncoupling of the DNA Topoisomerase and Replication Activities of an Initiator Protein

The replication initiator proteins encoded by the pT181 and related plasmids have sequence-specific DNA binding and topoisomerase activities. These proteins create a site-specific nick in one strand of the DNA at the origin of replication that serves as a primer for the initiation of replication. To...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1992-04, Vol.89 (7), p.3083-3087
Main Authors: Dempsey, Laurie A., Birch, Patrick, Khan, Saleem A.
Format: Article
Language:English
Subjects:
Amino acids
Bacterial Proteins - chemistry
Bacterial Proteins - genetics
Base Sequence
Binding Sites
Biochemistry
Biological and medical sciences
Deoxyribonucleic acid
DNA
DNA Replication
DNA Topoisomerases, Type I - chemistry
DNA Topoisomerases, Type I - metabolism
DNA-Binding Proteins - chemistry
DNA-Binding Proteins - metabolism
Electrophoresis
Fundamental and applied biological sciences. Psychology
Gels
Genetic Complementation Test
Medical research
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutant proteins
Nucleotides
Oligodeoxyribonucleotides - chemistry
Plasmids
Proteins
Recombinant Proteins - metabolism
Replication
Replication origin
Staphylococcus aureus
Structure-Activity Relationship
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Summary:The replication initiator proteins encoded by the pT181 and related plasmids have sequence-specific DNA binding and topoisomerase activities. These proteins create a site-specific nick in one strand of the DNA at the origin of replication that serves as a primer for the initiation of replication. To define the regions of the pT181-encoded initiator protein, RepC, that are involved in its DNA binding, topoisomerase, and replication activities, we have carried out site-directed mutagenesis of the repC gene. Analysis of mutant RepC proteins in vitro and in vivo has identified the amino acids that are critical for its various biochemical activities. The DNA binding domain of RepC was found to be located near its C-terminal region and was different from the domain involved in its sequence-specific topoisomerase activity. These studies also showed that the DNA topoisomerase activity of the initiator protein can be uncoupled from its tight noncovalent DNA binding and replication activites.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.89.7.3083