Available student projects

Macromolecular X-ray crystallography (MX) is the leading method for atomic-resolution structure determination in biology. Structure and dynamics of macromolecules determine their function, so MX provides mechanistic insights into life-enabling biochemical processes like photosynthesis, all our senses, the molecular basis of infection and disease, and structure-based pharmaceutical drug discovery [1]. The new MX3 beamline at the Australian Synchrotron promises to provide the high flux, microfocus beam required to push the frontiers of MX to static and time-resolved studies of tiny microcrystals of weakly-diffracting and/or radiation-sensitive macromolecules at room temperature (not currently possible in Australia [2]). This project aims to test the limits of MX3 capabilities through optimising serial macromolecular crystallography (SMX) by exploring the influence of experimental parameters on data quality and room temperature radiation damage (e.g. X-ray energy, bandwidth, focus size, exposure time/crystal, and various sample delivery approaches: standard goniometer, fixed target, in-tray screening, high-viscosity extrusion). The project involves X-ray physics, protein crystallography experiments, structural biology, high-performance computing, and close collaboration with the MX3 team [3]. 

[1] Pearson & Mehrabi, Curr. Op. Struct. Bio. 2020, 65:168-174.

[2] Martin-Garcia et al. IUCrJ (2017). 4, 439-454, and J. Synch. Rad. (2022), 29(3), 896-907.

[3] https://www.ansto.gov.au/high-performance-macromolecular-crystallography-beamline#content-capabilities 

Time-resolved serial macromolecular crystallography (TR-SMX) is a recently invented technique for direct visualisation of biomolecules in action, aka experimental “molecular movies” with atomic resolution. X-ray free-electron laser (XFEL) serial crystallography enables femtosecond-scale dynamics to be imaged in light-activated biomolecules [1], while reactions initiated by chemical binding are (currently) limited by microfluidic mixing and diffusion rates to ~ ms time scales [2].

This project aims to enable & apply time-resolved serial crystallography to image the dynamics of disulfide bond formation and small molecule binding in an enzyme involved in protein folding: disulphide bond-forming enzyme A (DsbA). DsbA is a key target for a new type of antibacterial drug to fight antimicrobial resistance, which is increasing worldwide [3]. However, DsbA is particularly sensitive to X-ray induced radiation damage at room temperature, so pursuing TR-SMX on DsbA requires a thorough understanding of local and global radiation damage. This project will investigate radiation damage in DsbA microcrystals, and compare room temperature structures from serial crystallography at a synchrotron vs an XFEL, where “diffraction before destruction” outruns structural X-ray-induced damage by using extremely brilliant femtosecond-scale X-ray pulses [1,2]. The project involves X-ray physics, serial crystallography experiments, structural biology, high-performance computing, close collaboration with Prof Begoña Heras’ lab (La Trobe University, [3,4]) and might include opportunities for experiments overseas. Once established, we will use TR-SMX to study DsbA interactions with small molecule inhibitors, substrates, and other enzymes to aid structure-based drug discovery based on DsbA inhibition [4]).

[1] Tenboer, Basu, Zatsepin et al. 2014. Science 346 (6214), 1242-1246.

[2] Stagno et al. 2017. Nature 541(7636), 242-246.

[3] Heras et al. 2009. Nature Reviews Microbiology 7 (3), 215-225.

[4] Smith, Paxman, Scanlon & Heras 2016. Molecules 21 (7), 811.

The ASU Compact X-ray Light Source (CXLS) is a novel, compact, hard X-ray source being constructed at Arizona State University based on inverse Compton scattering [1]. The CXLS aims to deliver synchrotron undulator-like capabilities on a table-top scale, with pulsed hard X-rays (100’s fs in duration at 1kHz), with a widely tuneable beam that will be usable for ultrafast spectroscopy, micro-crystallography and phase contrast medical imaging. CXLS will also comprise phase I of the construction of a recently-funded room-sized Compact X-ray free-electron laser (CXFEL) [2], and both are being considered as potential next-generation X-ray sources to build in Australia.

In this simulation project you will simulate serial crystallography data (X-ray diffraction from protein microcrystals with stochastically varying size, orientation, mosaicity) using nanoBragg [3]  to (a) explore the capabilities of CXLS for serial macromolecular crystallography (SMX), and (b) compare the performance of CrystFEL [4] and Careless [5] on difficult data (e.g. beam divergence, polychromaticity & crystal mosaicity and weak diffraction, limited detector dynamic range). CrystFEL is the most widely used suite of programs for serial crystallography data analysis [4], while Careless is a new tool for merging crystallography data that uses deep learning and variational inference [5]. The project is in collaboration with the CXLS/CXFEL team at ASU. This work will contribute to a pipeline for planning future experiments at CXLS as well as a science case for Australia to pursue such powerful and flexible sources. You may also have an opportunity to participate in the world-first SMX experiments at CXLS.

[1] https://news.asu.edu/20230205-discoveries-firstofitskind-instrument-officially-ushers-new-era-xray-science

[2] Graves et al. “ASU Compact XFEL” 2017. Proc. 38th Int. FEL Conference; https://biodesign.asu.edu/cxfel/

[3] nanoBragg. https://bl831.als.lbl.gov/~jamesh/nanoBragg/ and Sauter et al. 2020, Acta Cryst D 76(2), 176-192.

[4] White et al., Zatsepin & Chapman. 2013 Acta Cryst D. 69 (7), 1231-1240.

[5] Dalton, Greisman & Hekstra. 2022. Nature Comm. 13 (7764), 1-13.