Widespread but small-scale changes in the structural and dynamic properties of vaccinia virus poly(A) polymerase upon association with its processivity factor in solution.

Vaccinia virus poly(A) polymerase (VP55) has been analyzed via hydrogen-deuterium exchange (HDX) mass spectrometry in the absence and presence of its processivity factor, VP39, to improve our understanding of the mechanism by which processivity is impressed on the polymerase. For 119 peptic peptides covering 74.1% of VP55, the extent of HDX at 900 s was interpreted in the context of parameters deduced from the VP55-VP39 X-ray crystal structure. While HDX exhibited a degree of correlation with the mean SASA of whole residues within each peptide segment, HDX was generally more active than expected from either the SASA or hydrogen bonding status of the exchangeable amide proton, indicating a significant molecular dynamics contribution to amide proton deprotection. Peptic peptides undergoing either more or less HDX than expected were distributed throughout VP55 and showed consistency between multiple overlapping peptides. VP39 had a net, marginal cooling effect on VP55, indicating a possible restriction of VP55's flexibility. VP39's cooling effect was most extensive within the central domain of VP55's three domains, while a patch within VP55's C-terminal domain showed an increased level of HDX in the presence of VP39. Langevin dynamics all-atom simulations of VP55 motions showed slower relaxation to equilibrium in the absence of VP39. At equilibrium, regions showing extremes of variation in simulated atomic fluctuation were localized within VP55's N- and C-terminal domains, and VP39 had a predominantly cooling effect on VP55. Broadly, across VP55's peptic peptides, a mild negative correlation was noted between the extent to which deuteration was more active than predicted from the structure and the amplitudes of the simulated atomic fluctuation and/or degree of disorder at equilibrium.