Carbon-13 NMR studies of chloroplast membranes: carbon-13 enrichment without 13C-13C couplings.
ABSTRACT A novel approach to carbon-13 (13C) enrichment of chloroplast membranes (and plant materials in general) is presented for 13C-nuclear magnetic resonance (13C-NMR) studies. The method minimizes the occurrence of spectral complications arising from 13C-13C couplings resulting from a statistical distribution of 13C within the molecule with low probability of encountering two 13C atoms adjacent to each other. This is achieved by growing the plants in light surrounded by an atmosphere containing 1/3rd 12CO2 and 2/3rd 13CO2, liberated by weak acid-treatment of a mixture of corresponding barium carbonate salts.
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Article: Carbon-13 NMR studies of chloroplast membranes: carbon-13 enrichment without 13C-13C couplings.
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ABSTRACT: A new and simple approach for the determination of the temperature of gel-to-liquid crystalline phase transitions (Tc) of biological (chloroplast) membrane lipids from 13C-NMR resonance intensities is proposed. The variation of intensity of a temperature-sensitive NMR resonance is monitored by recording the spectra of the sample at a range of temperatures. From such a series of spectra recorded at different temperatures, a temperature-insensitive resonance is located. Then the ratio of the intensity of the temperature-sensitive to the intensity of the temperature-insensitive resonance is calculated from each spectrum to even out the procedural error, if any. The values of this ratio at different temperatures, when plotted against sample temperature, shows a break at Tc as confirmed by spin label ESR studies.Journal of Biochemical and Biophysical Methods 04/1990; 20(4):353-6. DOI:10.1016/0165-022X(90)90097-V · 1.81 Impact Factor
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ABSTRACT: Spinach chloroplast membranes were studied by natural abundance carbon-13 nuclear magnetic resonance (13C-NMR) spectroscopy in their normal state and after heat denaturation of membrane proteins. The membrane proteins were denatured by raising the temperature of the sample to 67 degrees C for 5 minutes [YashRoy, R.C. (1991) J. Biochem. Biophys. Methods 22, 55-59]. Line-broadening of 13C-NMR resonances arising from the 1st (carbonyl), 7th, 9th and 12th carbon atom of fatty-acyl chains with reference to the carbonyl (C-1) group shows increased immobilization of lipid fatty-acyl chains at these locations, obviously caused by changes in interactions between membrane lipids and proteins upon heat denaturation of membrane proteins.Journal of Biochemical and Biophysical Methods 10/1991; 23(3):259-61. DOI:10.1016/0165-022X(91)90019-S · 1.81 Impact Factor
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ABSTRACT: Aqueous dispersions of lipids isolated from spinach chloroplast membranes were studied by electron microscopy after negative staining with phosphotungstic acid. Influence of low temperature (5°C for 24 h) was also investigated. It was observed that when contacted with water, these lipids, as such, formed multilamellar structures. Upon sonication, these multilamellar structures gave rise to a clear suspension of unilamellar vesicles varying in size (diameter) between 250 and 750 Å. When samples of sonicated unilamellar vesicles were stored at 5°C for 24 h or more, they revealed a variety of lipid aggregates including liposomes, cylindrical rods (about 100 Å wide and up to 3600 Å long), and spherical micellar structures (100-200 Å in diameter)—thus indicating phase separation of lipids.Journal of Biosciences 07/1990; 15(2):93-98. DOI:10.1007/BF02703373 · 1.94 Impact Factor