It may be getting warmer outside,
but we have tips for good ice

In our May 2017 newsletter, we highlighted four membrane protein structures which have used either fluorinated Octyl Maltoside (O310F) or fluorinated Fos-Choline-8 (F300F) to improve the vitrification process in the Cryo-EM structure determination workflow. Although the exact mechanism of how these fluorinated surfactants improve the vitrification and subsequent data collection of membrane protein samples is still being determined, several hypotheses include: preventing protein interactions with the air/water interface; aiding preferred orientation issues by altering distribution of protein molecules in the vitreous ice; and/or, improving the distribution of ice thickness across the grid(1-4).  
 
In this month’s newsletter, we wanted to update you by providing recent examples from the past two years of these fluorinated surfactants during the vitrification step of membrane protein samples. In the table below, we also included a few examples of other vitrification additives which have also been used, including: CHAPS, Octyl Glucoside, and DMSO.
 
PDB Release Date PDB Protein Additive
2020-01-08 6PWN MscS mechanosensitive channel 0.01% f-OM
2019-09-04 6KG7 Piezo2 mechanosensitive channel 0.65 mM f-FC8
2019-08-28 6QTI Nicotinamide nucleotide proton channel 0.05% CHAPS
2019-08-07 6R7L SecYEG translocon 0.2% f-OM
2019-02-06 6E0H TMEM16 scramblase 3 mM f-FC8
2018-12-19 6N3Q Sec protein-translocation channel complex 3 mM f-FC8
2018-11-07 6H3I Type 9 secretion system translocon 1.5 mM f-FC8 or 0.7 mM f-OM
2018-10-24 6DMR TRPV5 ion channel 3 mM f-FC8
2018-10-17 6D3R CFTR 3 mM f-FC8
2018-09-26 6HJR Influenza Hemagglutinin 2% Octyl Glucoside
2018-08-08 6FOO Ryanodine receptor 1 0.2% f-OM
2018-08-01 6CJQ SthK CNG Potassium channel 3 mM f-FC8
2018-05-23 5YX9 TRPC6 ion channel 0.5 mM f-OM
2018-01-31 6C0V P-Glycoprotein transporter ABCB1 3 mM f-FC8
2017-12-27 6B5V TRPV5 ion channel 3 mM f-FC8
2017-12-13 6BPQ TRPM8 channel 2% DMSO


Tools for Cryo-EM:

Anatrace and Molecular Dimensions are continually developing tools and reagents to support the membrane and soluble protein Cryo-EM workflow. These include fluorinated surfactants to improve vitrification, amphipols (A8-35, PMAL-C8, and NAPol), lipid mixes for nanodisc reconstitution, protein stability screens, grid boxes and grid box storage pucks, and more. Additionally, Molecular Dimensions is now an authorized distributor of Protochips C-flat Cryo-EM grids in the Americas, Europe, and Asia! C-Flat grids will be in stock at both our UK and US offices for immediate shipment. Check out our Cryo-EM workflow page to learn more about all the latest tools we have to support your Cryo-EM experiments!

 
 References:
  1. Glaeser, RM, et al. (2017) Biophys Rep 3(1), 1-7.
  2. Noble, AJ, et al. (2018) Nat Methods 15(10), 793-795.
  3. Drulyte, I et al. (2018) Acta Crystallogr D Struct Biol 74(Pt 6), 560-571.
  4. Chen, J, et al. (2019) J Struct Biol X Volume 1. DOI: 10.1016/j.yjsbx.2019.100005