Conformational switch-defective X174 internal scaffolding proteins kinetically trap assembly intermediates before...

Area of Science:

• Virology
• Structural Biology
• Biochemistry

Background:

• Scaffolding proteins are crucial for virus assembly, often acting via conformational switches.
• Rapid viral morphogenesis typically prevents isolation of early assembly intermediates.
• Previous studies defined conformational switches by comparing final structures.

Purpose of the Study:

• To analyze early morphogenesis and scaffolding protein function in X174 bacteriophage.
• To investigate the role of specific amino acid residues in scaffolding protein-coat protein interactions.
• To identify and characterize novel molecular defects in viral assembly pathways.

Main Methods:

• Generated amino acid substitutions in the C-terminal region of the X174 scaffolding protein.
• Biochemically characterized mutant assembly pathways.
• Analyzed assembly intermediates and identified suppressor mutations in the viral coat protein.

Main Results:

• Identified two classes of molecular defects: protein binding and a novel conformational switching phenotype.
• Conformational switch mutations kinetically trapped pre-procapsid assembly intermediates.
• Discovered common second-site suppressors in the viral coat protein, suggesting a fluid assembly pathway.

Conclusions:

• X174 morphogenesis allows isolation of early intermediates, facilitating detailed analysis of scaffolding protein function.
• A single, coat protein conformational switch induced by the scaffolding protein is proposed.
• Improper or incomplete conformational switches kinetically trap assembly intermediates, providing a novel phenotype for studying viral assembly.