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Nucleic Acids Research, 1992, Vol. 20, No. 21 5549-5554
© 1992

MOLECULAR BIOLOGY

DNA conformational change induced by the bacteriophage {Phi}29 connector

José M. Valpuesta+, Manuel Serrano, Luis E. Donate+, Lucía Herranz§ and José L. Carrascosa*

Centro de Biologí Molecular (CSIC-UAM), Universidad Autónoma de Madrid 28049 Madrid, Spain

* To whom correspondence should be addressed

Received September 8, 1992. Revised October 15, 1992. Accepted October 15, 1992.

Translocation of viral DNA inwards and outwards of the capsid of double-stranded DNA bacteriophages occurs through the connector, a key viral structure that is known to interact with DNA. It is shown here that phage {Phi}29 connector binds both linear and circular double-stranded DNA. However, DNA-mediated protection of {Phi}29 connectors against Staphylococcus aureus endoprotease V8 digestion suggests that binding to linear DNA is more stable than to circular DNA. Endoprotease V8-protection assays also suggest that the length of the linear DNA required to produce a stable {Phi}29 connector-DNA interaction is, at least, twice longer than the {Phi}29 connector channel. This result is confirmed by experiments of {Phi}29 connector-protection of DNA against DNase I digestion. Furthermore, DNA circularization assays indicate that {Phi}29 connectors restrain negative supercoiling when bound to linear DNA. This DNA conformational change is not observed upon binding to circular DNA and it could reflect the existence of some left-handed DNA coiling or DNA untwisting inside of the {Phi}29 connector channel.

+Present addresses: Centro National de Biotecnología (CSIC), Universidad Autónoma de Madrid, 28049 Madrid

§Merck, Sharp & Dohme, Josefa Valcárcel 38, Madrid, Spain


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