Single-particle cryo-EM structures from iDPC-STEM at near-atomic resolution

Lazic, Ivan; Wirix, Maarten; Leidl, Max Leo; Haas, Felix de; Mann, Daniel; Beckers, Maximilian; Pechnikova, Evgeniya V.; Müller-Caspary, Knut; Egoavil, Ricardo; Bosch, Eric G. T. and Sachse, Carsten (2022): Single-particle cryo-EM structures from iDPC-STEM at near-atomic resolution. In: Nature Methods, Vol. 19, No. 9: pp. 1126-1136

Abstract

This paper explores the use of scanning transmission electron microscopy (STEM) to vitrified biological samples for biomolecular structure elucidation. Integrated differential phase contrast (iDPC)-STEM imaging of keyhole limpet hemocyanin and tobacco mosaic virus enabled cryo-EM structure determination at 6.5 and 3.5 angstrom resolution, respectively. In electron cryomicroscopy (cryo-EM), molecular images of vitrified biological samples are obtained by conventional transmission microscopy (CTEM) using large underfocuses and subsequently computationally combined into a high-resolution three-dimensional structure. Here, we apply scanning transmission electron microscopy (STEM) using the integrated differential phase contrast mode also known as iDPC-STEM to two cryo-EM test specimens, keyhole limpet hemocyanin (KLH) and tobacco mosaic virus (TMV). The micrographs show complete contrast transfer to high resolution and enable the cryo-EM structure determination for KLH at 6.5 angstrom resolution, as well as for TMV at 3.5 angstrom resolution using single-particle reconstruction methods, which share identical features with maps obtained by CTEM of a previously acquired same-sized TMV data set. These data show that STEM imaging in general, and in particular the iDPC-STEM approach, can be applied to vitrified single-particle specimens to determine near-atomic resolution cryo-EM structures of biological macromolecules.

• Item Type: Journal article
• Faculties: Chemistry and Pharmacy > Department of Chemistry
• Subjects: 500 Science > 540 Chemistry
• ISSN: 1548-7091
• Language: English