Spin-labeled nanobodies as protein conformational reporters for electron paramagnetic resonance in cellular membranes.

ABSTRACT: Nanobodies are emerging tools in a variety of fields such as structural biology, cell imaging, and drug discovery. Here we pioneer the use of their spin-labeled variants as reporters of conformational dynamics of membrane proteins using DEER spectroscopy. At the example of the bacterial ABC transporter TM287/288, we show that two gadolinium-labeled nanobodies allow us to quantify, via analysis of the modulation depth of DEER traces, the fraction of transporters adopting the outward-facing state under different experimental conditions. Additionally, we quantitatively follow the interconversion from the outward- to the inward-facing state in the conformational ensemble under ATP turnover conditions. We finally show that the specificity of the nanobodies for the target protein allows the dire

SUBMITTER: Galazzo L 

PROVIDER: S-EPMC7007536 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature