In situ distance measurements in a membrane transporter using maleimide functionalized orthogonal spin labels and 5-pulse electron double resonance spectroscopy

Spectroscopic investigation of membrane proteins in their native environment is a challenging task. Earlier we demonstrated the feasibility to measure precise distances within outer membrane proteins in E. coli and native membranes using methanethiosulfonate (MTS) functionalized labels combined with pulsed electron double resonance spectroscopy. Here we show the application of maleimide functionalized Gd(III), nitroxide, and trityl labels for in situ distance measurement using the cobalamin transporter BtuB. These labels enabled distance measurements for BtuB in E. coli and native outer membranes and in the membranes maleimide-Gd-DOTA also is effective. Further, we show that the observable dipolar evolution time can be significantly prolonged in the native environments using the Carr-Purcell 5-pulse electron double resonance sequence. For a nitroxide-nitroxide pair, application of sech/tanh inversion pulses substantially suppressed the 4-pulse artifact at the Q- band frequency. In the case of a nitroxide-trityl pair, Gaussian pump pulses of varying amplitude are sufficient to suppress the artifact to the typical noise level. The feasibility of a range of bioresistant spin labels and the 5-pulse electron double resonance offers promising tools for investigating heterooligomeric membrane protein complexes in their native environment.

Validation of Structural Grounds for Anomalous Molecular Mobility in Ionic Liquid Glasses

Ionic liquid (IL) glasses have recently drawn much interest as unusual media with unique physicochemical properties. In particular, anomalous suppression of molecular mobility in imidazolium IL glasses vs. increasing temperature was evidenced by pulse Electron Paramagnetic Resonance (EPR) spectroscopy. Although such behavior has been proven to originate from dynamics of alkyl chains of IL cations, the role of electron spin relaxation induced by surrounding protons still remains unclear. In this work we synthesized two deuterated imidazolium-based ILs to reduce electron–nuclear couplings between radical probe and alkyl chains of IL, and investigated molecular mobility in these glasses. The obtained trends were found closely similar for deuterated and protonated analogs, thus excluding the relaxation-induced artifacts and reliably demonstrating structural grounds of the observed anomalies in heterogeneous IL glasses.

Transient morphology of lithium anodes in batteries monitored by in operando pulse electron paramagnetic resonance

Enhancing lithium-ion battery technology in terms of specific capacity and charging time is key for the advancement of the electrification of transportation. Particularly for fast charging, inhomogeneous deposition of metallic lithium, for example on commercial graphite or metallic lithium anodes, leads to cell degradation and safety issues. To understand the underlying mechanisms and develop counter-measures, non-invasive online detection techniques providing satisfactory time resolution are crucial. Here, we demonstrate in operando pulse electron paramagnetic resonance to observe transient processes during pulsed fast charging in cells with metallic lithium anodes. Sampling timescales of 100 ms enable real-time monitoring of the formation and evolution of porous lithium during and after charging pulses. It was observed that the generated morphology continued to evolve after the end of a charging pulse, whereby surface features were fusing with a time constant that was slower than their formation.