No abstract

Microscopic redox equilibrium constants, a new species-specific type of physicochemical parameters, were introduced and determined to quantify thiol-disulfide equilibria of biological significance. The thiol-disulfide redox equilibria of glutathione with cysteamine, cysteine, and homocysteine were approached from both sides, and the equilibrium mixtures were analyzed by quantitative NMR methods to characterize the highly composite, co-dependent acid-base and redox equilibria. The directly obtained, pH-dependent, conditional constants were then decomposed by a new evaluation method, resulting in pH-independent, microscopic redox equilibrium constants for the first time. The 80 different, microscopic redox equilibrium constant values show close correlation with the respective thiolate basicities and provide sound means for the development of potent agents against oxidative stress.

The combination of 19F NMR spectroscopy tagging and paramagnetic relaxation enhancement (PRE) NMR spectroscopy experiments was evaluated as a versatile method to probe protein–protein interactions and conformational changes of intrinsically disordered proteins upon complex formation. The feasibility of the approach is illustrated with an application to the Myc‐Max protein complex; this is an oncogenic transcription factor that binds enhancer box DNA fragments. The single cysteine residue of Myc was tagged with highly fluorinated [19F]3,5‐bis(trifluoromethyl)benzyl bromide. Structural dynamics of the protein complex were monitored through intermolecular PREs between 19F‐Myc and paramagnetic (1‐oxyl‐2,2,5,5‐tetramethyl‐Δ3‐pyrroline‐3‐methyl)methanethiosulfonate (MTSL)‐tagged) Max. The 19F R2 relaxation rates obtained with three differently MTSL‐tagged Max mutants revealed novel insights into the differential structural dynamics of Myc‐Max bound to DNA and the tumour suppressor breast cancer antigen 1. Given its ease of implementation, fruitful applications of this strategy to structural biology and inhibitor screening can be envisaged.