Solution Wide-angle X-ray Scattering (waxs) and its Application to Envelope Based Phasing

Solution X-ray scattering from biological macromolecules is an increasingly important technique that yields low-resolution structural information on the molecules, and can be used to monitor their shape and conformational changes (1) ,(2), (3). In contrast to numerous successful applications of small angle X-ray scattering (SAXS), e.g. ref. (4-6), practical use of wide-angle X-ray scattering (WAXS) data has been limited due to the weak scattering despite of the higher-resolution structural information. Recently, we have succeeded in measuring accurate WAXS data on protein solutions at a standard macromolecular diffraction station (7). SAXS data were collected at a dedicated SAXS station, and combined with WAXS data to give a full scattering curve out to 2.5 A resolution. Both indirect and direct Fourier transforms of the full scattering pattern exhibit that some high resolution aspects of the structural hierarchy and function of a protein can be investigated in solution.To elucidate the importance of solution WAXS data, we have employed the high resolution scattering curve to solve the phase problem, which remains central to crystallographic structure determination. A 6-dimensional search method of molecular replacement (FSEARCH) was used to locate a low-resolution molecular envelope within the crystallographic unit cell. The most probable model selected from a dozen bead models constructed at 5 A resolution was employed to phase the single-crystal diffraction data. We find that inclusion of WAXS data is essential for correctly locating the molecular envelope in the crystal unit cell, as well as for locating heavy atom sites (8). The initial phases can be used as a starting point for a variety of phase-extension techniques; successful application of which will result in complete phasing of a crystallographic data set and determination of a macromolecule's internal structure to atomic resolution.