DNA packaging by lambda-like bacteriophages: mutations broadening the packaging specificity of terminase, the...

Area of Science:

• Molecular Biology
• Virology
• Genetics

Background:

• Bacteriophages lambda and 21 exhibit distinct DNA packaging specificities.
• These specificities are mediated by the small terminase subunits, which possess DNA-binding motifs like the recognition helix.

Purpose of the Study:

• To investigate the molecular basis of DNA packaging specificity in phages lambda and 21.
• To elucidate the role of specific DNA-binding motifs in terminase subunits for host DNA recognition and packaging.

Main Methods:

• Construction of chimeric terminase subunits by swapping recognition helix domains.
• Analysis of DNA packaging efficiency and plaque formation of mutant phages.
• Identification and characterization of suppressor mutations affecting terminase-DNA interactions.

Main Results:

• A chimeric terminase with the lambda-terminase backbone and 21-terminase recognition helix preferentially packaged 21 DNA, with a 20-fold reduction in lambda DNA packaging.
• Pseudorevertant mutations in the recognition helix or the cosB binding site altered DNA packaging specificity.
• Mutations in the recognition helix either disrupted unfavorable DNA contacts or created new nonspecific contacts, leading to loss of discrimination.
• Mutations in cosB altered base-pair specificity, enhancing packaging by 21-specific terminase and reducing it by lambda-terminase.

Conclusions:

• The recognition helix and specific DNA contacts within the cosB site are critical for determining DNA packaging specificity in phages lambda and 21.
• The support helix and turn are essential for robust packaging discrimination, while the wing motif does not contribute to cosB specificity.
• These findings support a model of evolutionary divergence from a common ancestor with broad packaging specificity.