Molecular dynamics of proteins investigated by NMR relaxation methods

The nuclear magnetic resonance relaxation times of solvent water nuclei are known to decrease upon addition of diamagnetic solute protein. For this reason NMR relaxation methods are able to provide information on molecular dynamics changes of water protons and their interaction with macromolecules' surfaces. We present results of measurements of relaxation rates R1 = 1/T1, R2 = 1/T2 and R1ρ = 1/T1ρ in the rotating frame for three proteins: chicken egg white lysozyme, egg white albumin, and bovine serum albumin, obtained at proton resonant frequency of 60 MHz. Besides the relaxation rates dependences on concentration in the 4-23% (g/100 g solution) range, the analysis of the Carr-Purcell-Meiboom-Gill CPMG multi-echo T2 experiments with variable pulse rate τ was performed. The dependences of relaxation rates on protein concentration are linear at low concentration. When protein concentration increases the slope of the straight line rapidly changes at so-called "critical" concentration which depends on MW of the diluted protein. Investigated dispersion of T2, obtained using the CPMG method with a variable pulse rate, for concentrations higher and lower than the "critical" one, exhibits unequal behavior. At high concentration one-exponential curves and at low concentration two-exponential curves correspond closely with experimental data. The obtained parameters of exponents allow an estimation of the ratio of the amount of water with the determined motion freedom, that is free and bounded water, in solution. We showed that the CPMG dispersion method applied to aqueous protein solutions may widen the current understanding of the nature of molecular dynamics of hydrated water protons in non-perturbed environment

NMR relaxation measurements as a tool for observation of oxidative processes

Water proton relaxation times, T₁ and T₂, were measured to assess the kinetics of the oxidative processes in biological samples. The oxidation in aqueous solutions of albumins was promoted by an addition of 3% hydrogen peroxide (H₂O₂). Immediately following this addition a sharp exponential decrease of both relaxation times was observed. As we confirmed experimentally, the time course of relaxation depended on several essential factors like structure and the concentration of proteins and also the presence of antioxidants added to solution. In experiments with protein solutions containing a small amount of ascorbic acid, after reaching a minimum, relaxation time increased towards the initial (pre-addition H₂O₂) values. We conclude that this T₁ and T₂ recovery is a consequence of the presence of antioxidants and may be used to evaluate its action. This study demonstrates that nuclear magnetic resonance (NMR) relaxation measurements may be useful in evaluating free radicals reactions and antioxidants capacity

Investigation of starch hydration by 2D time domain NMR

Proton exchange between spin groups of the solid matrix of hydrated granular potato starch and water was studied using the 2D time domain NMR. The proton spin-spin relaxation time T$\text{}_{2}$, and spin-lattice relaxation time T$\text{}_{1}$ (selective and non-selective pulse sequences) were measured at room temperature. The observed spin relaxation results were analysed for exchange assuming a two-site exchange model (between water and solid matrix of starch). In this analysis we determined the intrinsic spin-lattice relaxation time for water protons (49 ms) and solid starch matrix protons (172 ms), as well as the water-starch magnetization exchange rate (86 s$\text{}^{-1}$)

NMR Relaxation Measurements as a Tool for Observation of Oxidative Processes

Water proton relaxation times, T₁ and T₂, were measured to assess the kinetics of the oxidative processes in biological samples. The oxidation in aqueous solutions of albumins was promoted by an addition of 3% hydrogen peroxide (H₂O₂). Immediately following this addition a sharp exponential decrease of both relaxation times was observed. As we confirmed experimentally, the time course of relaxation depended on several essential factors like structure and the concentration of proteins and also the presence of antioxidants added to solution. In experiments with protein solutions containing a small amount of ascorbic acid, after reaching a minimum, relaxation time increased towards the initial (pre-addition H₂O₂) values. We conclude that this T₁ and T₂ recovery is a consequence of the presence of antioxidants and may be used to evaluate its action. This study demonstrates that nuclear magnetic resonance (NMR) relaxation measurements may be useful in evaluating free radicals reactions and antioxidants capacity

Magnetic resonance assays of haloperidol in human serum albumin

Haloperidol is a commonly used neuroleptic drug associated with a range of side effects. The aim of the study was to determine in vitro interaction of haloperidol with human serum albumin (HSA) by 19F magnetic resonance spectroscopy (MRS) at 9.4 Tesla (T). The practical measurement based on fluorine resonance has been proposed to determine drug level in HSA at 37\ub0C.Peer reviewed: YesNRC publication: Ye