A Small-Angle X-ray Scattering Apparatus for Studying Biological Macromolecules in Solution

This paper describes the development of a simple laboratory-based small-angle X-ray scattering apparatus for the study of biological macromolecules in solution. The instrument is based on a two-circular-aperture collimation design combined with a conventional rotating-anode Cu Ka X-ray source, a graphite monochromator and a multiwire area detector. The geometry of the collimator, the beam-stop-to-detector distance and the thickness of the platinum foil of the defining aperture have been optimized to reduce background scattering. The effective Q range is from 0.01 to 0.33 Å⁻¹, where Q = (4π sin θ)/λ is the magnitude of the scattering vector, 2θ is the scattering angle and λ is the wavelength of the X-rays. The length of the collimator, the pinhole sizes and the helium-flushed sample-to-detector path can be easily changed depending on the resolution and intensity requirements of an experiment. The diffraction pattern of a polycrystalline pellet of ammonium sulfate mounted about 2.5 cm in front of the beam stop and 40 cm in front of the detector is used to monitor changes in the incident-beam intensity as well as the differences in absorption of X-rays by the sample solutions and the solvents, to ensure correct background subtractions. Data collection is controlled by a computer through a parallel DMA (direct memory access) I/O module. Data collection and reduction software has been developed. The typical data collection time is about 2 h for a 5 mg ml⁻¹ 10 kDa protein dissolved in an aqueous solution. Examples of applications of this small-angle X-ray scattering instrument to studying protein size and conformation changes are presented.