microfluidic-chips-for-situ-crystal-x-ray-diffraction-situ-dynamic

Microfluidic Chips for <em>In Situ</em> Crystal X-ray Diffraction and <em>In Situ</em> Dynamic Light Scattering for Serial Crystallography

This protocol describes fabricating microfluidic devices with low X-ray background optimized for goniometer based fixed target serial crystallography. The ...

University of Hamburg

Many biochemical processes take place on timescales ranging from femto-seconds to seconds. Accordingly, any time-resolved experiment must be matched to the speed of the structural changes of interest. Therefore, the timescale of interest defines the requirements of the X-ray source, instrumentation and data-collection strategy. In this study, a minimalistic approach for  crystallization is presented that requires only a few microlitres of sample solution containing a few hundred crystals. It is demonstrated that complete diffraction data sets, merged from multiple crystals, can be recorded within only a few minutes of beamtime and allow high-resolution structural information of high quality to be obtained with a temporal resolution of 40 ms. Global and site-specific radiation damage can be avoided by limiting the maximal dose per crystal to 400 kGy. Moreover, analysis of the data collected at higher doses allows the time-resolved observation of site-specific radiation damage. Therefore, our approach is well suited to observe structural changes and possibly enzymatic reactions in the low-millisecond regime.

Svetlana Kapis

Institute for Biochemistry and Molecular Biology,
Laboratory for Structural Biology of Infection and Inflammation,
Institute for Biochemistry and Molecular Biology, Laboratory for Structural Biology of Infection and Inflammation

University of Hamburg

Microfluidic Chips for In Situ Crystal X-ray Diffraction and In Situ Dynamic Light Scattering for Serial Crystallography