High-speed atomic force microscopy (HS-AFM [1]) is a powerful technique that provides dynamic movies of biomolecules at work. HS-AFM has the notable advantage that it permits to subject the proteins under investigation to environmental cues such as changes of pH, ions, ligands, temperature, light and force. This is particularly advantageous for the study of ion channels that respond to a wide range of stimuli [2], and transporters with substrate-dependent activities. To break current temporal limitations to characterize molecular dynamics using HS-AFM, we use HS-AFM line scanning (HS-AFM-LS) and HS-AFM height spectroscopy (HS-AFM-HS), techniques whereby we reduce the image acquisition dimension to single scan lines and a fixed position where the oscillating HS-AFM tip detects the motions of the molecules under the tip [3]. These methods allowed us to determine the dynamics of individual unlabeled amino acid transporters at millisecond temporal resolution [4].
Top: HS-AFM line scanning kymograph with 3.3ms temporal resolution of GltPh transport motion. Bottom: Height/time trace and idealized state assignment (red).
References:
[1] A high-speed atomic force microscope for studying biological macromolecules, PNAS, 2001, 98(22):12468-12472, doi.org/10.1073/pnas.211400898, Toshio Ando, Noriyuki Kodera, Eisuke Takai, Daisuke Maruyama, Kiwamu Saito, and Akitoshi Toda
[2] Force-induced conformational changes in Piezo1, Nature, 2019, 573(7773):230–234, doi. 10.1038/s41586-019-1499-2, Yi-Chih Lin, Yusong R Guo, Atsushi Miyagi, Jesper Levring, Roderick MacKinnon & Simon Scheuring
[3] High-Speed AFM Height Spectroscopy (HS-AFM-HS): Microsecond dynamics of unlabeled biomolecules, Nature Communications, 2018, 9(1):4983. doi:10.1038/s41467-018-07512-3, George Heath & Simon Scheuring
[4] Millisecond dynamics of an unlabeled amino acid transporter, Nature Communications, 2020, 11(5016), doi.org/10.1038/s41467-020-18811-z, Tina R Matin, George R Heath, Gerard HM Huysmans, Olga Boudker & Simon Scheuring
https://umontreal.zoom.us/j/4074103908?pwd=NUNpcXVueGRMTFVhdHRvZFpvaFNDdz09
Identifiant: 407 410 3908
Mot de passe: trois quatre trois six un un un
