Cryo-SAXS method for solution structure from nanoliter volumes

Figure 1. Cryo-SAXS reconstructions. (a) A cryo-cooled droplet containing macromolecules in solution. (b-d) Cryo-SAXS shape reconstructions over-layed with their atomic structures: glucose isomerase (b), lysozyme(c), and 25bp DNA (d).

Small angle X-ray scatering (SAXS) is a powerful technique for measuring the structure of biomolecules and their interactions in solution. However, SAXS requires large volumes of sample to compensate for radiation sensitivity of macromolecules. To overcome the radiation damage limit, we adapted a technique from X-ray crystallography, where solutions are rapidly cooled to temperatures of ~100K. For robust background subtraction, I designed a windowless sample holder and protocol for measuring and compensating for variable sample thickness [3]. With these innovations, we demonstrated the first shape reconstructions of proteins and DNA from frozen droplets (Fig. 1) [1]. In addition, I contributed to the design of a microfabricated sample holder for high-throughput applications [2].


Research Articles

[1]

Meisburger SP, Warkentin M, Chen H, Hopkins JB, Gillilan RE, Pollack L, Thorne RE. 2013. Breaking the Radiation Damage Limit with Cryo-SAXS. Biophys J 104, 227-36. PMID: 23332075, DOI: 10.1016/j.bpj.2012.11.3817

[2]

Hopkins JB, Katz AM, Meisburger SP, Warkentin MA, Thorne RE, Pollack L. 2015. A microfabricated fixed path length silicon sample holder improves background subtraction for cryoSAXS. J Appl Cryst 48, 227-37. PMID: 26089749, DOI: 10.1107/S1600576714027782

Patents

[3] Meisburger SP, Warkentin MA, Hopkins JB, Katz AM, Pollack L, and Thorne RE. Apparatus and methods for low temperature small angle X-ray scattering. Dec 2013. WO Patent App. PCT/US2013/044,088.