NTP binding induces conformational changes in the double-stranded RNA bacteriophage ø6 subviral particles

Area of Science:

• Molecular Virology
• Structural Biology
• Biochemistry

Background:

• Bacteriophage ø6 is a double-stranded RNA virus with a nucleocapsid (NC) and surrounding membrane.
• The ø6 RNA polymerase complex, comprising proteins P1, P2, P4, and P7, is located beneath the major coat protein P8.
• Protein P1 forms the dodecahedral framework for the polymerase complex.

Purpose of the Study:

• To develop a novel method for isolating stable bacteriophage ø6 RNA polymerase complex particles.
• To investigate the stabilizing effects of purine nucleotides on the polymerase complex.
• To analyze nucleotide-induced conformational changes within the viral nucleocapsid.

Main Methods:

• Development of a new isolation technique for stable polymerase complex particles.
• Assessment of particle stability and polymerase activity over time.
• Use of monoclonal antibodies to detect conformational changes in nucleocapsid proteins upon nucleotide binding.

Main Results:

• Stable polymerase complex particles were successfully isolated, retaining integrity and activity for several days.
• Purine nucleotides, particularly GTP, dGTP, ddGTP, and GDP, demonstrated efficient stabilization of the particles.
• Nucleotide binding induced conformational changes in the nucleocapsid, altering antibody epitope accessibility for proteins P4, P8, and potentially P1.

Conclusions:

• A robust method for isolating stable bacteriophage ø6 RNA polymerase complexes has been established.
• Nucleotide binding is crucial for stabilizing the polymerase complex and induces significant structural rearrangements in the nucleocapsid.
• These findings provide insights into the structural dynamics of bacteriophage RNA polymerase and its interaction with the nucleocapsid.