Figure 4
From: Protein delivery to living cells by thermal stimulation for biophysical investigation
(A) Model structure of human ubiquitin protein labelled with Maleimide DOTA Gadolinium tag in the S20C and G35C position derived from MMM44. (B) Nitroxide double integral signal decay over time extracted from the RT cw-EPR X-band spectra for the Ubiquitin S20C/G35C ma-Px delivered cells. The spectra DI (Double Integral) were plotted against the overall time course. The first point time point take into account the death time before the sample acquisition composed by the delivery and preparation time. Each cw-EPR experiment was set to be ~ 13 min in acquisition time. The asterisks represent the relative time position in which the pulsed EPR samples were frozen. (C) Acquired EDFS spectra for the delivered cells, recorded at 50 K at the center of the resonator dip under overcoupling conditions. The cellular Manganese (Mn2+) EDFS signal is contributing to the spectra with the broad transition and the downfield peak in respect to the nitroxide signal. (D) DEER traces of the in vitro (blue trace) and in-cell samples (cyan traces) acquired at 50 K.
