The use of a mini-κ goniometer head in macromolecular crystallography diffraction experiments

Abstract

Most macromolecular crystallography (MX) diffraction experiments at synchrotrons use a single-axis goniometer. This markedly contrasts with small-molecule crystallography, in which the majority of the diffraction data are collected using multi-axis goniometers. A novel miniaturized κ-goniometer head, the MK3, has been developed to allow macromolecular crystals to be aligned. It is available on the majority of the structural biology beamlines at the ESRF, as well as elsewhere. In addition, the Strategy for the Alignment of Crystals (STAC) software package has been developed to facilitate the use of the MK3 and other similar devices. Use of the MK3 and STAC is streamlined by their incorporation into online analysis tools such as EDNA. The current use of STAC and MK3 on the MX beamlines at the ESRF is discussed. It is shown that the alignment of macromolecular crystals can result in improved diffraction data quality compared with data obtained from randomly aligned crystals.

Keywords: crystal alignment; data-collection strategies; kappa goniometer.

Figure 1

The EMBL/ESRF mini-κ goniometer head (MK3) as mounted on the MD2M diffractometer on ID14-4 in a ‘closed’ and in a fully ‘open’ conformation, highlighting the three rotational axes and the α angle.

Figure 2

A schematic representation of STAC. The core framework, which can manage input or output information flow (yellow) from/to various external software solutions, is shown in magenta. The GUI interface, with the various ‘tabs’, is shown in blue. The service modules for alignment and strategy calculation as well as for hardware access are shown in green.

Figure 3

Screenshot of STAC, illustrating how it can be used in either a standalone or a server application mode, such as through the workflow tab in MxCuBE. The STAC GUI shows the ‘Input for Re-Orientation’ tab with the ‘Anomalous Data Collection’ options for a P2₁2₁2₁ crystal system.

Figure 4

SHELXC results highlighting the increase in the anomalous signal that can be achieved by measuring an aligned crystal along a twofold axis: (a) for a high-energy (E = 13.2 keV) bovine trypsin S-SAD phasing experiment, (b) for an Se-SAD phasing experiment.

Figure 5

CC_all versus CC_weak SHELXD plots for the three data sets in Table 3 ▶: (a) c* -aligned data set, (b) non-aligned data set, (c) merged data set.

Figure 6

A plot of the axial reflections along a*, b* and c* highlighting how most of the systematic absences can be experimentally measured by collecting data at different orientations.

Figure 7

Plot of the maximal oscillation width possible to avoid overlapping spots as calculated using BEST (Bourenkov & Popov, 2010 ▶) (a) for a deliberately misaligned thaumatin test crystal at 1.4 Å resolution and (b) for a 70S ribosome test crystal at 3.8 Å resolution.