Comparison of membrane organization in mitochondria from yeast and rat liver by nuclear magnetic resonance spectroscopy

Abstract

Proton magnetic resonance (PMR) and carbon-13 magnetic resonance (CMR) spectra of intact, unsonicated yeast and rat liver motochondria show differences which may be correlated with the composition of the membranes. High resolution PMR and CMR signals in intact yeast mitochondria have been assigned to regions of fluid lipid-lipid interaction on the basis of spectra of extracted lipid and protein, and the temperature dependence of NMR signals from the intact membrane. PMR spectra suggest that about 20% of total yeast phospholipid is in regions where both intramolecular fatty acid chain mobility and lateral diffusion of entire phospholipid molecules are possible. No such regions apear to exist in rat liver mitochondria. For both yeast and rat liver mitochondria, comparison of PMR and CMR spectra suggests that about 50% of phospholipid appears to be in regions where intramolecular fatty acid chain motion is considerable, but lateral diffusion is restricted. The remaining phospholipid appears to have little inter- or intramolecular mobility. Since NMR observation of lipid extracts from membranes indicates that phospholipid-sterol interactions do not account for the spectra of intact mitochondria, these effects are interpreted in terms of extensive lipid-protein interactions.

Proton magnetic resonance spectroscopy of promitochondrial membranes from yeast grown under different regimes of lipid supplementation

“…Spectra of the three types of promitochondria were obtained either immediately after preparation and washing in 0.2 M KCI in D20, or after lyophilizing and resuspending in DeO, as detailed previously (Brown et al, 1975). The differences in spectra were minor for the same type of promitochondria subjected to these two different preparative procedures.…”

“…The strain of Saccharomyces cerevisiae used, and the general growth conditions have been described previously (Gordon & Stewart, 1971 ;Brown et al, 1975). Where indicated, the growth medium was supplemented with ergosterol or cholesterol (30 rag/liter) and/or unsaturated fatty acid in the form of Tween 80 (polyoxyethylene sorbitan monoleate, 5 ml/ liter); Tween 20 (polyoxyethylene sorbitan monolaurate) was substituted for Tween 80 when unsaturated fatty acid supplementation was not required.…”

“…Lipids were extracted from promitochondria and assayed as described previously (Brown et al, 1975). Two-dimensional thin-layer chromatography was carried out as described by Getz, Jakovcic, Heywood, Frank and Rabinowitz (1970), except that the plates were not washed with diethyl ether between development in the first and second dimensions, since this step tended to remove material from the surface of the plates.…”

“…Promitochondria were prepared from cell homogenates using the same methods described previously for preparation of mitochondria (Brown et al, 1975).…”

“…In the preceding paper (Brown, Bradbury, Austin & Stewart, 1975), we reported that mitochondria from aerobically grown yeast yield high resolution nuclear magnetic resonance spectra. The results indicate that the NMR spectra obtained reflect the mobility of a certain fraction of the lipid components of membranes.…”