Use of paramagnetic 19F NMR to monitor domain movement in a glutamate transporter homolog.
 

Use of paramagnetic 19F NMR to monitor domain movement in a glutamate transporter homolog.

http://www.bionmr.com//www.ncbi.nlm....s-npg_logo.gif Related Articles Use of paramagnetic 19F NMR to monitor domain movement in a glutamate transporter homolog.

Nat Chem Biol. 2020 Jun 08;:

Authors: Huang Y, Wang X, Lv G, Razavi AM, Huysmans GHM, Weinstein H, Bracken C, Eliezer D, Boudker O

Abstract
In proteins where conformational changes are functionally important, the number of accessible states and their dynamics are often difficult to establish. Here we describe a novel 19F-NMR spectroscopy approach to probe dynamics of large membrane proteins. We labeled a glutamate transporter homolog with a 19F probe via cysteine chemistry and with a Ni2+ ion via chelation by a di-histidine motif. We used distance-dependent enhancement of the longitudinal relaxation of 19F nuclei by the paramagnetic metal to assign the observed resonances. We identified one inward- and two outward-facing states of the transporter, in which the substrate-binding site is near the extracellular and intracellular solutions, respectively. We then resolved the structure of the unanticipated second outward-facing state by cryo-EM. Finally, we showed that the rates of the conformational exchange are accessible from measurements of the metal-enhanced longitudinal relaxation of 19F nuclei.


PMID: 32514183 [PubMed - as supplied by publisher]



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