Erythrosin B as a New Photoswitchable Spin Label for Light-Induced Pulsed EPR Dipolar Spectroscopy

We present a new photoswitchable spin label for light-induced pulsed electron paramagnetic resonance dipolar spectroscopy (LiPDS), the photoexcited triplet state of erythrosin B (EB), which is ideal for biological applications. With this label, we perform an in-depth study of the orientational effects in dipolar traces acquired using the refocused laser-induced magnetic dipole technique to obtain information on the distance and relative orientation between the EB and nitroxide labels in a rigid model peptide, in good agreement with density functional theory predictions. Additionally, we show that these orientational effects can be averaged to enable an orientation-independent analysis to determine the distance distribution. Furthermore, we demonstrate the feasibility of these experiments above liquid nitrogen temperatures, removing the need for expensive liquid helium or cryogen-free cryostats. The variety of choices in photoswitchable spin labels and the affordability of the experiments are critical for LiPDS to become a widespread methodology in structural biology.

Automated summary

In this study, we introduce a new photoswitchable spin label that can be used to investigate the orientational effects in pulsed electron paramagnetic resonance dipolar spectroscopy. Furthermore, experiments conducted at temperatures above liquid nitrogen eliminate the need for expensive cryogenic equipment.