No observable non-thermal effect of microwave radiation on the growth of microtubules.

Despite widespread public interest in the health impact of exposure to microwave radiation, studies of the influence of microwave radiation on biological samples are often inconclusive or contradictory. Here we examine the influence of microwave radiation of frequencies 3.5 GHz, 20 GHz and 29 GHz on the growth of microtubules, which are biological nanotubes that perform diverse functions in eukaryotic cells. Since microtubules are highly polar and can extend several micrometres in length, they are predicted to be sensitive to non-ionizing radiation. Moreover, it has been speculated that tubulin dimers within microtubules might rapidly toggle between different conformations, potentially participating in computational or other cooperative processes. Our data show that exposure to microwave radiation yields a microtubule growth curve that is distorted relative to control studies utilizing a homogeneous temperature jump. However, this apparent effect of non-ionizing radiation is reproduced by control experiments using an infrared laser or hot air to heat the sample and thereby mimic the thermal history of samples exposed to microwaves. As such, no non-thermal effects of microwave radiation on microtubule growth can be assigned. Our results highlight the need for appropriate control experiments in biophysical studies that may impact on the sphere of public interest.

Bacteriorhodopsin: Structural Insights Revealed Using X-Ray Lasers and Synchrotron Radiation.

Directional transport of protons across an energy transducing membrane-proton pumping-is ubiquitous in biology. Bacteriorhodopsin (bR) is a light-driven proton pump that is activated by a buried all-trans retinal chromophore being photoisomerized to a 13-cis conformation. The mechanism by which photoisomerization initiates directional proton transport against a proton concentration gradient has been studied by a myriad of biochemical, biophysical, and structural techniques. X-ray free electron lasers (XFELs) have created new opportunities to probe the structural dynamics of bR at room temperature on timescales from femtoseconds to milliseconds using time-resolved serial femtosecond crystallography (TR-SFX). Wereview these recent developments and highlight where XFEL studies reveal new details concerning the structural mechanism of retinal photoisomerization and proton pumping. We also discuss the extent to which these insights were anticipated by earlier intermediate trapping studies using synchrotron radiation. TR-SFX will open up the field for dynamical studies of other proteins that are not naturally light-sensitive.

Lipidic cubic phase serial millisecond crystallography using synchrotron radiation.

Lipidic cubic phases (LCPs) have emerged as successful matrixes for the crystallization of membrane proteins. Moreover, the viscous LCP also provides a highly effective delivery medium for serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs). Here, the adaptation of this technology to perform serial millisecond crystallography (SMX) at more widely available synchrotron microfocus beamlines is described. Compared with conventional microcrystallography, LCP-SMX eliminates the need for difficult handling of individual crystals and allows for data collection at room temperature. The technology is demonstrated by solving a structure of the light-driven proton-pump bacteriorhodopsin (bR) at a resolution of 2.4 Å. The room-temperature structure of bR is very similar to previous cryogenic structures but shows small yet distinct differences in the retinal ligand and proton-transfer pathway.

Natively inhibited Trypanosoma brucei cathepsin B structure determined by using an X-ray laser.

The Trypanosoma brucei cysteine protease cathepsin B (TbCatB), which is involved in host protein degradation, is a promising target to develop new treatments against sleeping sickness, a fatal disease caused by this protozoan parasite. The structure of the mature, active form of TbCatB has so far not provided sufficient information for the design of a safe and specific drug against T. brucei. By combining two recent innovations, in vivo crystallization and serial femtosecond crystallography, we obtained the room-temperature 2.1 angstrom resolution structure of the fully glycosylated precursor complex of TbCatB. The structure reveals the mechanism of native TbCatB inhibition and demonstrates that new biomolecular information can be obtained by the "diffraction-before-destruction" approach of x-ray free-electron lasers from hundreds of thousands of individual microcrystals.

Membrane protein crystallization from lipidic phases.

Membrane protein structural biology is enjoying a steady acceleration in the rate of success. Nevertheless, numerous membrane protein targets are resistant to the traditional approach of directly crystallizing detergent solubilized and purified protein and the 'niche market' of lipidic phase crystallization is emerging as a powerful complement. These approaches, including lipidic cubic phase, lipidic sponge phase, and bicelle crystallization methods, all immerse purified membrane protein within a lipid rich matrix before crystallization. This environment is hypothesized to contribute to the protein's long-term structural stability and thereby favor crystallization. Spectacular recent successes include the high-resolution structures of the beta(2)-adrenergic G-protein-coupled receptor, the A(2A) adenosine G-protein-coupled receptor, and the mitochondrial voltage dependent anion channel. In combination with technical innovations aiming to popularize these methods, lipidic phase crystallization approaches can be expected to deliver an increasing scientific impact as the field develops.