Supercoiled DNA wraps around the bacteriophage phi 29 head-tail connector.

Area of Science:

• Molecular Biology
• Virology
• Structural Biology

Background:

• The bacteriophage phi 29 is a model system for studying viral DNA packaging.
• The head-tail connector is a critical component of the phi 29 DNA packaging machinery.
• Understanding DNA packaging mechanisms is crucial for viral replication and gene therapy applications.

Purpose of the Study:

• To investigate the interaction between the isolated Bacillus subtilis bacteriophage phi 29 head-tail connector and supercoiled DNA.
• To elucidate the role of the connector in the initiation phase of DNA packaging.
• To characterize the structural and functional aspects of DNA wrapping by the connector.

Main Methods:

• Electron microscopy (EM) for determining DNA contour length and visualizing complexes.
• Scanning transmission EM for mass and radial density determinations.
• Topoisomerase I treatment to assess DNA supercoil restraint.

Main Results:

• Supercoiled pBR322 DNA wraps around the exterior of the phi 29 head-tail connector.
• DNA contour length decreased by ~180 base pairs per bound connector.
• Increased complex mass indicated ~170 base pairs of DNA bound externally.
• Connectors restrained DNA supercoils and inefficiently bound non-supercoiled DNA.

Conclusions:

• The wrapping of supercoiled DNA by the isolated phi 29 connector mimics the initiation of viral DNA packaging.
• This interaction involves DNA wrapping and supercoil restraint, preceding linearization and translocation.
• The findings provide insights into the mechanism of DNA packaging in bacteriophages and viruses.