Modeling truncated pixel values of faint reflections in MicroED images

Abstract

The weak pixel counts surrounding the Bragg spots in a diffraction image are important for establishing a model of the background underneath the peak and estimating the reliability of the integrated intensities. Under certain circumstances, particularly with equipment not optimized for low-intensity measurements, these pixel values may be corrupted by corrections applied to the raw image. This can lead to truncation of low pixel counts, resulting in anomalies in the integrated Bragg intensities, such as systematically higher signal-to-noise ratios. A correction for this effect can be approximated by a three-parameter lognormal distribution fitted to the weakly positive-valued pixels at similar scattering angles. The procedure is validated by the improved refinement of an atomic model against structure factor amplitudes derived from corrected microelectron diffraction (MicroED) images.

Ab initio structure determination from prion nanocrystals at atomic resolution by MicroED

“…When high-resolution intensities collected using MicroED are combined with phases calculated from preexisting models by the method of molecular replacement, they result in atomic resolution structures (19)(20)(21)(22). However, to date, no other X-ray crystallographic method for experimental phase determination has been adapted to MicroED.…”

Atomic-resolution structures from fragmented protein crystals with the cryoEM method MicroED

“…Proteinase K ( E. album ) crystals were grown by combining 2 µL of protein solution (50 mg/ml) with 2 µL of precipitant solution (1.0–1.3 M ammonium sulfate, 0.1 M Tris pH 8.0) 10 .…”

“…Sweeps were corrected to account for negative pixel values 10 , then indexed and integrated in MOSFLM 27 using its graphical interface iMOSFLM 28 , or in XDS 29 . To further probe the scattering power of the crystallites, diffraction patterns were initially processed without imposing any resolution cutoffs other than those entailed by the geometry of the experiment.…”

“…Electrons interact strongly with matter so that large crystals are not required to detect high-resolution Bragg reflections 8 . MicroED has been used to determine the structures of the enzymes lysozyme 5,9 , proteinase K 10 , catalase 11,12 , and Ca 2+ -ATPase 12 . It was also recently used to solve the 1.4 Å resolution structure of the toxic NAC-core of α-synuclein, where diffraction data were collected from crystals smaller than the wavelength of visible light 13 .…”

Protein structure determination by electron diffraction using a single three-dimensional nanocrystal

“…A quantitative comparison between image plates and a Medipix hybrid pixel detector indicated a substantial improvement to be offered by the latter (Georgieva et al, 2011;Nederlof et al, 2013). Hybrid pixel detectors such as Medipix, Timepix and EIGER (Llopart et al, 2002Johnson et al, 2012) are well suited for measuring high-energy electrons (McMullan et al, 2007), and can overcome difficulties in detecting weak peaks, for example for CCD and CMOS detectors (Hattne et al, 2016;Rodriguez & Gonen, 2016). …”