NMR relaxivity changes in chloroplast suspensions. Effects of NH2OH and of treatments altering the redox state of the photosynthetic electron transport chain

Abstract

Treatments (illumination, chemical oxidation or reduction) which are potentially capable of producing paramagnetic centers in chloroplast thylakoid membranes do not produce enhancements of the proton magnetic relaxivities of these preparations. However, exposure of thylakoid membranes to varying concentrations of hydroxylamine induces a time-dependent increase in relaxivity for which the steady-state magnitude is dependent on hydroxylamine concentration. The appearance of relaxivity is correlated kinetically with inactivation of oxygen-evolving centers; in addition both processes show a threshold effect with respect to hydroxylamine concentration. Kinetic analyses of these hydroxylamine-induced effects suggest that at low (less than or equal to 100 microM) and at intermediate (200--500 microM) concentrations, hydroxylamine extraction is partially counteracted by a reverse process that reactivates oxygen-evolving centers in the dark.

Effects of physical and chemical treatments on chloroplast manganese. NMR and ESR studies

“…The experiments of Robinson et al [29] did not extend to high enough [NH2OH ] for R~ to exhibit the maximum we found in R 2. Also, it is noteworthy that they observed R z in chloroplast suspensions treated with 0.5 and 1.5 mM TPB (at comparable [Chl]) and found no effect.…”

“…Recently, Robinson et al [29] reported the effects of up to 1 mM NH2OH on the proton R I (spin-lattice) or chloroplast suspensions. Their independent results, while qualitatively similar to ours, are shifted to 10-fold higher [NH2OH ].…”

Topology of NH2OH-induced Mn(II) release from chloroplast thylakoid membranes

“…Apparatus for the NMR measurements (all at 3°C, 20.7 MHz) has been described previously [26,27]. Preillumination of thylakoids was carried out at 25 -+ I°C using a coil of clear-walled catheter tubing as previously described [31].…”

“…Blankenship and Sauer [25] exploited this approach, utilizing the characteristic 6-line ESR spectrum of hexaquomanganese produced by Tris treatment to show that this Mn(II) is apparently sequestered by thylakoid membranes at an EDTA-accessible site. An alternative method for monitoring Mn(II) topology is provided by the enhancement of the nuclear magnetic relaxation rate of water protons observed after NH2OH inactivation of water oxidation [26,27]. In constrast to the ESR experiment, which monitors only the soluble hexaquomanganese species, NMR measurements of spin lattice relaxation rates (TI -l =R1)are sensitive primarily to the presence of membrane-bound Mn(II) (or strictly, to Mn(II) bound in sites of long reorientational correlation time [26]).…”

“…We have utilized the NMR experiment in extensive investigations to analyze the origins of R 1 in thylakoid suspensions [26][27][28][29][30][31] and to identify the causes of light-induced changes in R1 [28,30]. We have in addition utilized this technique to monitor the effects of NH2OH on water oxidation and to explore the mechanism by which NH2OH inactivates this reaction [31 ].…”