Three macromolecular crystallography (MX) beamlines at the Helmholtz-Zentrum Berlin (HZB) are available for the regional, national and international structural biology user community. The state-of-the-art synchrotron beamlines for MX BL14.1, BL14.2 and BL14.3 are located within the low-β section of the BESSY II electron storage ring. All beamlines are fed from a superconducting 7 T wavelength-shifter insertion device. BL14.1 and BL14.2 are energy tunable in the range 5-16 keV, while BL14.3 is a fixed-energy side station operated at 13.8 keV. All three beamlines are equipped with CCD detectors. BL14.1 and BL14.2 are in regular user operation providing about 200 beam days per year and about 600 user shifts to approximately 50 research groups across Europe. BL14.3 has initially been used as a test facility and was brought into regular user mode operation during the year 2010. BL14.1 has recently been upgraded with a microdiffractometer including a mini-κ goniometer and an automated sample changer. Additional user facilities include office space adjacent to the beamlines, a sample preparation laboratory, a biology laboratory (safety level 1) and high-end computing resources. In this article the instrumentation of the beamlines is described, and a summary of the experimental possibilities of the beamlines and the provided ancillary equipment for the user community is given.

Massive fragment screening effort provides a foundation for the development of macrodomain inhibitors as novel antiviral agents.

The last decade witnessed a tremendous growth in combined efforts of biologists, chemists and physicists to understand the dominant factors determining the specificity and directionality of transmembrane transfer processes in proteins. A large variety of experimental techniques is being used including X-ray and neutron diffraction, but also time-resolved optical, infrared and magnetic resonance spectroscopy. This is done in conjunction with genetic engineering strategies to construct site-specific mutants for controlled modification of the proteins. As a general perception of these efforts, the substantial influence of weak interactions within the protein and its membrane interfaces is recognized. The weak interactions are subject to subtle changes during the reaction cycle owing to the inherent flexibility of the protein-membrane complex. Specific conformational changes accomplish molecular-switch functions for the transfer process to proceed with optimum efficiency. Characteristic examples of time varying non-bonded interactions are specific H-patterns and/or polarity effects of the microenvironment. The present perception has emerged from the coupling of newly developed spectroscopic techniques – and advanced EPR certainly deserves credit in this respect – with newly developed computational strategies to interpret the experimental data in terms of protein structure and dynamics. By now, the partners of this coupling, particularly high-field EPR spectroscopy and DFT-based quantum theory, have reached a level of sophistication that applications to large biocomplexes are within reach. In this review, a few large paradigm biosystems are surveyed which were explored lately in our laboratory. Taking advantage of the improved spectral and temporal resolution of high-frequency/high-field EPR at 95 GHz/3.4 T and 360 GHz/12.9 T, as compared to conventional X-band EPR (9.5 GHz/0.34 T), three biosystems are characterized with respect to structure and dynamics: (1) Light-induced electron-transfer intermediates in wild-type and mutant reaction-centre proteins from the photosynthetic bacterium Rhodobacter sphaeroides, (2) light-driven proton-transfer intermediates of site-specifically nitroxide spin-labelled mutants of bacteriorhodopsin proteins from Halobacterium salinarium, (3) refolding intermediates of site-specifically nitroxide spin-labelled mutants of the channel-forming protein domain of Colicin A bacterial toxin produced in Escherichia coli. The detailed information obtained is complementary to that of protein crystallography, solid-state NMR, infrared and optical spectroscopy techniques. A unique strength of high-field EPR is particularly noteworthy: it can provide highly desired detailed information on transient intermediates of proteins in biological action. They can be observed and characterized while staying in their working states on biologically relevant time scales. The review introduces the audience to origins and basic experiments of EPR in relation to NMR, describes the underlying strategies for extending conventional EPR to high-field/high-frequency EPR, and highlights those details of molecular information that are obtained from high-field EPR in conjunction with genetic engineering and that are not accessible by “classical” spectroscopy. The importance of quantum-chemical interpretation of the experimental data by DFT and advanced semiempirical molecular-orbital theory is emphasized. A short description of the laboratory-built 95 GHz and 360 GHz EPR/ENDOR spectrometers at FU Berlin is also presented. The review concludes with an outlook to future opportunities and challenges of advanced bio-EPR in interdisciplinary research.

We present design details of and first measurements with a novel continuous wave (cw) high-field/high-frequency electron paramagnetic resonance spectrometer operating at a microwave frequency of 360 GHz and a magnetic field of up to 14 T. The spectrometer design incorporates a heterodyne mixer detection scheme with a quasi-optical transmission line and a bimodal induction mode Fabry–Perot cavity. First cw experiments on polycrystalline 1,1-diphenyl-2-picryl-hydrazyl and bisdiphenylene-β-phenylallyl benzolate in polystyrene at room temperature and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl in frozen solution at 190 K demonstrate the high Zeeman resolution achievable and allow an estimate of the present detection sensitivity of 4×109 spins/G at a detection bandwidth of 1 Hz.

Two new macromolecular crystallography (MX) beamlines at the National Synchrotron Light Source II, FMX and AMX, opened for general user operation in February 2017 [Schneider et al. (2013). J. Phys. Conf. Ser. 425, 012003; Fuchs et al. (2014). J. Phys. Conf. Ser. 493, 012021; Fuchs et al. (2016). AIP Conf. Proc. SRI2015, 1741, 030006]. FMX, the micro-focusing Frontier MX beamline in sector 17-ID-2 at NSLS-II, covers a 5–30 keV photon energy range and delivers a flux of 4.0 × 10¹² photons s⁻¹ at 1 Å into a 1 µm × 1.5 µm to 10 µm × 10 µm (V × H) variable focus, expected to reach 5 × 10¹² photons s⁻¹ at final storage-ring current. This flux density surpasses most MX beamlines by nearly two orders of magnitude. The high brightness and microbeam capability of FMX are focused on solving difficult crystallographic challenges. The beamline's flexible design supports a wide range of structure determination methods – serial crystallography on micrometre-sized crystals, raster optimization of diffraction from inhomogeneous crystals, high-resolution data collection from large-unit-cell crystals, room-temperature data collection for crystals that are difficult to freeze and for studying conformational dynamics, and fully automated data collection for sample-screening and ligand-binding studies. FMX's high dose rate reduces data collection times for applications like serial crystallography to minutes rather than hours. With associated sample lifetimes as short as a few milliseconds, new rapid sample-delivery methods have been implemented, such as an ultra-high-speed high-precision piezo scanner goniometer [Gao et al. (2018). J. Synchrotron Rad. 25, 1362–1370], new microcrystal-optimized micromesh well sample holders [Guo et al. (2018). IUCrJ, 5, 238–246] and highly viscous media injectors [Weierstall et al. (2014). Nat. Commun. 5, 3309]. The new beamline pushes the frontier of synchrotron crystallography and enables users to determine structures from difficult-to-crystallize targets like membrane proteins, using previously intractable crystals of a few micrometres in size, and to obtain quality structures from irregular larger crystals.