Supplementary Fig. 3: RP-HPLC of the TEMPO-HOCbl preparation with detection at 316 nm. | Nature Protocols

Supplementary Fig. 3: RP-HPLC of the TEMPO-HOCbl preparation with detection at 316 nm.

From: In situ observation of conformational dynamics and protein ligand–substrate interactions in outer-membrane proteins with DEER/PELDOR spectroscopy

Supplementary Fig. 3

The major peak is highlighted and an overlay of its UV-Vis spectra with the same for cyanocobalamin is shown as an inset on the left. This peak revealed a strong EPR signal. The shoulder on the left exhibited a weak signal, while the other small peaks gave no EPR signal (data not shown). Comparison of the EPR spectrum of the crude mixture (50 μM as estimated from A316, in magenta in the inset on the right) with a 100 μM 4-amino TEMPO standard (in green) revealed 65±10% spin content.

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