Long Distance Measurements up to 160 A in the GroEL Tetradecamer Using Q-Band DEER EPR Spectroscopy.

ABSTRACT: Current distance measurements between spin-labels on multimeric protonated proteins using double electron-electron resonance (DEER) EPR spectroscopy are generally limited to the 15-60 Å range. Here we show how DEER experiments can be extended to dipolar evolution times of ca. 80 μs, permitting distances up to 170 Å to be accessed in multimeric proteins. The method relies on sparse spin-labeling, supplemented by deuteration of protein and solvent, to minimize the deleterious impact of multispin effects and substantially increase the apparent spin-label phase memory relaxation time, complemented by high sensitivity afforded by measurements at Q-band. We demonstrate the approach using the tetradecameric molecular machine GroEL as an example. Two engineered surface-exposed mutants, R268C and E

SUBMITTER: Schmidt T 

PROVIDER: S-EPMC5166617 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature