6WX6

Cryo-EM Structure of Human Apoferritin Light Chain Vitrified Using Back-it-up

Summary for 6WX6

• Entry DOI: 10.2210/pdb6wx6/pdb
• EMDB information: 21951
• Descriptor: Ferritin light chain, CALCIUM ION (3 entities in total)
• Functional Keywords: human, apoferritin, ferritin, light chain, back-it-up, through-grid wicking, metal binding protein
• Biological source: Homo sapiens (Human)
• Total number of polymer chains: 24
• Total formula weight: 582942.18

Authors

Tan, Y.Z.,Rubinstein, J.L. (deposition date: 2020-05-09, release date: 2020-05-20, Last modification date: 2024-03-06)

Primary citation

Tan, Y.Z.,Rubinstein, J.L.
Through-grid wicking enables high-speed cryoEM specimen preparation.
Acta Crystallogr D Struct Biol, 76:1092-1103, 2020

PubMed Abstract: Blotting times for conventional cryoEM specimen preparation complicate time-resolved studies and lead to some specimens adopting preferred orientations or denaturing at the air-water interface. Here, it is shown that solution sprayed onto one side of a holey cryoEM grid can be wicked through the grid by a glass-fiber filter held against the opposite side, often called the `back', of the grid, producing a film suitable for vitrification. This process can be completed in tens of milliseconds. Ultrasonic specimen application and through-grid wicking were combined in a high-speed specimen-preparation device that was named `Back-it-up' or BIU. The high liquid-absorption capacity of the glass fiber compared with self-wicking grids makes the method relatively insensitive to the amount of sample applied. Consequently, through-grid wicking produces large areas of ice that are suitable for cryoEM for both soluble and detergent-solubilized protein complexes. The speed of the device increases the number of views for a specimen that suffers from preferred orientations.

PubMed: 33135680
DOI: 10.1107/S2059798320012474

Experimental method

ELECTRON MICROSCOPY (2 Å)