J. Bacteriol., Sep 1996, 5222-5228, Vol 178, No. 17
Copyright © 1996, American Society for Microbiology

An 18-base-pair sequence is sufficient for termination of rolling- circle replication of plasmid pT181

AC Zhao and SA Khan
Department of Molecular Genetics and Biochemistry, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.

pT181 and related plasmids of gram-positive bacteria replicate by a rolling-circle mechanism. The replication initiator protein of pT181, RepC, has origin-specific nicking-closing activities. Replication of the plasmid pT181 leading strand initiates by covalent extension of the RepC-generated nick, and the origin of replication contains signals for both initiation and termination of DNA replication. We have investigated the sequence requirements for the initiation and termination steps by using plasmids containing two pT181 origins. In vitro replication experiments showed that 18- and 24-bp synthetic oligonucleotides containing the RepC nick site were active in the termination of replication. However, initiation of replication required a larger region which also includes the RepC binding site. Plasmids containing the 18- and 24-bp region were also found to be nicked by the RepC protein. Our results demonstrate that sequence requirements for initiation and termination of pT181 replication overlap, but while the RepC binding site is required for initiation, it is dispensable for termination.

This article has been cited by other articles:

• Zhao, A. C., Ansari, R. A., Schmidt, M. C., Khan, S. A. (1998). An Oligonucleotide Inhibits Oligomerization of a Rolling Circle Initiator Protein at the pT181 Origin of Replication. J. Biol. Chem. 273: 16082-16089 [Abstract] [Full Text]  
• del Solar, G., Giraldo, R., Ruiz-Echevarria, M. J., Espinosa, M., Diaz-Orejas, R. (1998). Replication and Control of Circular Bacterial Plasmids. Microbiol. Mol. Biol. Rev. 62: 434-464 [Abstract] [Full Text]