Search all library resources

Developing angular intensity correlations of X-ray photons as a tool for studying structures of proteins in non-crystalline solutions

This dissertation offers lessons learned and tools developed as we attempt to apply correlated X-ray scattering (CXS) to non-crystalline proteins in solution. It builds on our previous work of extracting correlation signals from scattering intensities of ensembles of mental nanoparticles, which has led to three-dimensional (3D) structural insights not reflected in azimuthally averaged measurements. In his 1977 paper, Zvi Kam proposed the idea of correlating X-ray photons scattered by an ensemble of randomly oriented particles suspended in solution. He found that if the exposure time is much shorter than the diffusion timescale of Brownian motion, correlations between photons scattered into different angles encode 3D structural information of the particles not accessible via conventional small or wide-angle X-ray scattering. The advent of the X-ray free electron laser (XFEL) renders Kam's idea feasible for non-crystalline solutions of proteins. With femtosecond pulses and extremely high fluences, the XFEL is not only capable of probing ensembles of molecules essentially frozen in time but also delivering a large number of photons per pulse, a capability critical for enhancing angular intensity correlation signals. Meanwhile, probing proteins in solution removes the need for crystallization, allows measurements of mixtures of conformational states under physiological conditions, and broadens opportunities for time-resolved experiments. The body of work in this dissertation draws from scattering data collected with samples containing the G-protein Gi alpha subunit during two separate beam times conducted at the Linac Coherent Light Source. The Gi alpha subunit was chosen for these proof-of-principle experiments because of its important role in the G-protein coupled receptor signaling pathway. This dissertation has taken the first steps in developing and validating CXS as a tool for probing ensembles of biomolecules in solution. These first steps as well as ideas described in this dissertation to improve CXS towards a mature pipeline that yields reliable and detailed structural insights aim to inspire others in the solution scattering community to engage with the unique challenges and rewards of this technique.