(499.11) Differences in Conformational Dynamics Between a Viral Polyprotein and Its Processed Products in Functionally Relevant Regions Revealed by Solution-State NMR Spectroscopy

Poster Board Number: A213

Dennis Winston (The Pennsylvania State University), David Boehr (The Pennsylvania State University)

Viruses have evolved numerous strategies to expand protein function despite their limited genetic material, including proteolytic cleavage of polyproteins to obtain proteins with different functions than their precursors. In this way, proteolysis enables temporal control over the viral life cycle by varying the amounts of proteins in competing cleavage pathways. The 72 kD poliovirus polyprotein 3CD is involved in important protein-protein, protein-RNA and protein-lipid interactions in viral replication and infection. It is a precursor to 3C protease and 3D RNA-dependent RNA polymerase, but has different protease specificity, is not an active polymerase, and participates in different protein and RNA interactions than its processed products. Despite the existence of X-ray crystal structures for the three proteins, the molecular mechanism for the different functionality of 3CD is not well understood due to the structure of the domains in 3CD being nearly identical to the isolated 3C and 3D proteins.

It has been proposed that despite nearly identical X-ray crystal structures, the conformational dynamics of 3CD in solution may be different than those of 3C and 3D, allowing 3CD to adopt additional conformations and perform new functions compared to its processed products. To this end, we compared the conformational dynamics of 3C, 3D, and 3CD using solution-state NMR spectroscopy. We used Carr-Purcell-Meiboom-Gill (CPMG) multiple quantum relaxation dispersion, methyl 1H chemical exchange saturation transfer (CEST), and methyl spin-spin relaxation to probe conformational dynamics from the picosecond to second timescales. We have identified differences in conformational dynamics in functionally important regions that may be useful in understanding the differences in function between 3C, 3D, and 3CD. In particular, expansion of the conformational ensemble available to 3CD may allow it to perform additional functions not seen in 3C and 3D alone despite having nearly identical lowest-energy structures.

Support or Funding Information

This work was partly funded through NIH grant AI104878z