Escherichia coli Tus protein acts to arrest the progression of DNA replication forks in vitro.
A polar DNA replication barrier is formed when the DNA-binding protein Tus forms a complex with any of the four 23-base-pair terminator (ter) sites found in the terminus region of the Escherichia coli chromosome. We have used a plasmid DNA replication system reconstituted with purified proteins in vitro to investigate the interaction of the Tus protein with the replication fork. Purified Tus protein alone is necessary and sufficient to arrest DNA replication on ColE1-type plasmid templates containing ter sites. Tus protein-catalyzed termination depends upon the orientation of the ter site in the plasmid DNA. Nucleotide resolution mapping of the terminated nascent DNA shows that leading-strand DNA synthesis arrests at the point of contact with the Tus protein, while the final lagging-strand primer sites are 50-70 nucleotides upstream. In addition, the distribution of leading-strand arrest sites changes when the composition of the proteins on the lagging-strand side of the replication fork is altered.
Hill
T M
TM
Department of Biosciences and Biotechnology, Drexel University, Philadelphia, PA 19104.
Marians
K J
KJ
D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D004261:DNA Replication; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins; D007700:Kinetics; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010957:Plasmids; D013728:Terminator Regions, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D015345:Oligonucleotide Probes; C061910:tus protein, Bacteria; C489236:tus protein, E coli