On-grid enrichment strategies in cryo-EM: affinity-functionalized supports for low-abundance and challenging targets

Affinity-functionalized and engineered cryo-EM supports are transforming specimen preparation by enabling on-grid enrichment and improved recovery of challenging biological assemblies. This review provides a critical assessment of current platforms, emerging technologies and the remaining challenges that must be addressed to establish affinity-based specimen preparation as a routine component of cryo-EM workflows.

A short history of Auto-Rickshaw

Auto-Rickshaw is an example of practical, decision-driven automated macromolecular structure determination from crystallographic diffraction data. This review traces its development at the EMBL Hamburg/DESY beamline environment, where rapid validation of diffraction experiments was essential, through its expansion into a general platform for automated structure solution and its continuing relevance to beamline automation.

The protein epiphany

Eleanor Dodson examines George Sheldrick's contributions to macromolecular crystallography.

Separating the effects of temperature and absorbed X-ray dose on unit-cell volume

The nature of the expansion of the crystal unit-cell volume with absorbed dose and with temperature was examined for three different proteins.

Radiation damage in serial femtosecond crystallography studied in hemoglobin

Protein structures determined by serial femtosecond crystallography are indistinguishable when measured with 3 and 10 fs X-ray pulses, indicating no structural benefit from further shortening the pulse duration under the studied conditions.

Perspectives for pharmaceutical screening at XFEL sources using the example of Lassa virus endonuclease

To facilitate structure-based drug development, XFEL sources are of paramount interest. Here, we analyse the feasibility and data quality of serial femtosecond X-ray crystallography compound screening at Linac Coherent Light Source and compare it with PETRA III synchrotron experiments. For one Lassa virus model protein, we determined new crystal structures with two inhibitors.

Single-crystal X-ray and in crystallo XANES studies resolve cobalt coordination in cobalamin and rule out radiation-induced ligand loss in human intrinsic factor

High-resolution crystallography combined with in crystallo Co K-edge XANES shows that cobalamin retains octahedral cobalt coordination in both oxidized and reduced states. These results rule out radiation-induced Co–CN cleavage as the cause of the missing axial ligand in intrinsic factor-bound cobalamin.

FPGA-based scanner and SerialEM server for 4D-STEM electron tomography

Software for a multipurpose hardware board enables flexible scanning with a ready acquisition and processing pipeline to implement 4D-STEM electron tomography.