HATSAXS : a versatile high-throughput system for solution SAXS

• Main Author: Lotze, Gudrun
• Published: University of Reading 2012
• Subjects: 543.59
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.606949

A new, very flexible setup has been developed for solution small-angle X-ray scattering (SAXS) for biomolecular solutions that is capable of measuring samples fully automatically. The proposed system is based on a multi droplet sample approach, where buffer and sample droplets separated by air gaps are loaded at the same time into fluorinated polymer tubing connected to a glass X-ray cell. The droplets are driven into the path of the X-ray beam by an automated syringe pump. This thesis demonstrates that such a system is capable of performing successful SAXS experiments on weakly scattering biological samples in a fully automated manner. It exhibits a high rate of reproducibility and meets the demand for an extremely accurate background subtraction. The risk of radiation damage is eliminated by a constant mono-directional or oscillating sample flow mode improving as well the data quality. The droplet size was reduced to a 1111 volume allowing for the successful investigation of very scarce biological samples, such as transmembrane proteins. Full automation was achieved by installing a droplet identification algorithm following a change in transmission over time. Potential cross-contamination effects for very high concentrated samples were identified, quantified, and reduced to a minimum. The system provides as well a highly mobile and adaptive concept already successfully operated at several beamlines that are being upgraded to support highly automated throughput SAXS (HATSAXS) experiments due to the incorporation of the here proposed system. Proof-of-concept data is presented on lysozyme and ovalbumin, together with a bioSAXS study on bifidobacterium bifidum j3-galactosidase, an enzyme of real biological interest . .