Time-resolved SAXS Data Research Articles

High Resolution Time-Resolved SAXS shows that RNA-Induced SV40 Capsid Protein Self-Assembly is very Fast and without Detectable Concentrations of Intermediates

Many spherical viruses assemble their capsids around their nucleic acid, a reaction that may involve hundreds of individual molecules. Yet, uniform virus-like particles appear to assemble quickly. To dissect the assembly mechanism and genome encapsidation, we have established a simplest case experimental system. We show that mixing SV40 VP1 pentamers with RNA 500 mers yields T=1 particles comprised of 12 pentamers and one RNA molecule. We examined the kinetics of this reaction by Time-Resolved Small Angle X-ray Scattering (TR-SAXS). TR-SAXS shows that assembly is very fast; the reaction is nearly third-complete at 35 ms when mixing 0.5 μM RNA and 7.5 μM VP1 pentamers. Nonetheless, assembly appears to be a two-state process with only free pentamers and capsids observed; intermediates are undetectable. Finally, we show that TR-SAXS data are very well fit by master equations that describe assembly as a nucleation of an RNA molecule with one, two or three pentamers, followed by a cascade of elongation reactions in which one pentamer is added at a time. From this model, we estimate that the molar rate constant for addition of pentamers is approximately 109 M−1 s−1. Such a rate is possible only if facilitated by long-ranged distance protein-nucleic acid attraction. We therefore suggest that the growing nucleo-protein complex is able to act as an electrostatic antenna for attracting other capsid subunits.

A Novel Approach to Extract Morphological Variables in Crystalline Polymers from Time-Resolved Synchrotron SAXS Data

A novel approach to extract morphological variables in crystalline polymers from time-resolved SAXS data using the method of correlation and interface distribution functions has been devised. The principle of the calculation is based on two alternative expressions of Porod's law using the form of the interference function. The approach enables the continuous estimate of the Porod constant, and corrections for liquid scattering and finite interface between the two phases, from the time-resolved data. A model of lamellar morphology has been implemented to interpret the calculated correlation and interface distribution functions. Many detailed morphological variables such as lamellar long period, thicknesses of crystal and amorphous phases, interface thickness, and scattering invariant can be estimated. An example analysis of isothermal crystallization in PET measured by synchrotron SAXS is demonstrated.