The long-term objective of this research program is to understand the structural and dynamic bases of protein recognition leading to the assembly of viruses. This knowledge will permit identification of steps along the assembly pathways that can be targeted for interference and control. The plan utilizes static and dynamic Raman spectroscopic probes in combination with complementary hydrodynamic and calorimetric methods to elucidate protein structures and interactions.
The specific aims of the proposed research are as follows: (1) Identify protein main-chain conformations, side-chain interactions and covalent modifications that regulate the assembly and function of the specialized portal vertices of icosahedral viruses. (2) Determine new structural information on the viral terminase complexes that function as ATP-fueled motors to package viral genomes within capsids. (3) Ascertain kinetic and thermodynamic controls of discrete steps in viral capsid morphogenesis, especially those relating to procapsid formation, capsid maturation and stabilization of the capsid/genome interface. (4) Establish new qualitative and quantitative correlations to advance the effectiveness of Raman spectroscopy as a diagnostic probe of viral protein structures and assembly pathways. The investigations will focus on key viruses for which complementary genetic and biochemical data are now available or obtainable, including isometric dsDNA and dsRNA virions (P22, HK97, PRD1, F6), filamentous virions (fd, Pf1, Pf3, PH75) and related subviral assemblies. The project will exploit novel probes of protein structure, dynamics and interactions within the framework of the virus assembly/disassembly paradigm. Experimental protocols will incorporate isotope-edited Raman (IER), near-infrared (NIR) laser Raman, ultraviolet-resonance Raman (UVRR) and polarized Raman spectroscopies, including temperature-dependent and H/D exchange-dependent applications. The plan is designed specifically to advance understanding of the assembly roles of viral subunit folding, structure, dynamics and recognition, and generally to expand the capabilities of laser Raman spectroscopy as a probe of biological complexes. The biological significance of the proposed research derives from the critical need for fundamental knowledge about the molecular basis of virus assembly in order to understand and control viral diseases.
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