James S. Vincent

Raman spectroscopic studies of dimyristoylphosphatidic acid and its interactions with ferricytochrome c in cationic binary and ternary lipid-protein complexes.

The vibrational Raman spectra of both pure 1-alpha-dimyristoylphosphatidic acid (DMPA) liposomes and DMPA multilayers reconstituted with ferricytochrome c at pH 7 and pH 4, with either sodium or calcium as the cation, are reported as a function of temperature. Multilayers composed of a 1:1 mol ratio DMPA and dimyristoylphosphatidylcholine with perdeuterated acyl chains (DMPC-d54) have also been reconstituted with approximately 10(-4) M ferricytochrome c for Raman spectroscopic observation. Total integrated band intensities and relative peak height intensity ratios, two spectral Raman scattering parameters used to characterize bilayer properties, are sensitive to the presence of both ferricytochrome c and the cation in the reconstituted liposomes. Temperature profiles, derived from the various Raman intensity parameters for the 3,100-2,800 cm-1 lipid acyl chain C-H stretching mode region specifically reflect bilayer perturbations due to the interactions of ferricytochrome c. At pH 4 the calcium DMPA multilamellar gel to liquid crystalline phase transition temperatures Tm, defined by either the C-H stretching mode I2850/I2880 and I2935/I2880 peak height intensity ratios, are 58.5 +/- 0.5 degrees C and 60.0 +/- 0.3 degrees C, respectively. This difference in Tms resolves the phase transition process into first an expansion of the lipid lattice and then a melting of the lipid acyl chains. At pH 7 the calcium DMPA liposomes show no distinct phase transition characteristics below 75 degrees C. For sodium DMPA liposomes reconstituted with ferricytochrome c at either pH 4.0 or pH 7.0, spontaneous Raman spectra show altered lipid structures at temperatures above 40 degrees C. Resonance Raman spectra indicate that ferricytochrome c reconstituted in either calcium or sodium DMPA liposomes changes irreversibly above Tm. For either the binary lipid or ternary lipid-protein systems reconstituted with DMPC-d54, linewidth parameters of the DMPC-d54 acyl chain CD2 symmetric stretching modes at 2,103 cm-1 provide a sensitive measure of the conformational and dynamic properties of the perdeuterated lipid component, while the 3,000 cm-1 C-H spectral region reflects the bilayer characteristics of the DMPA species in the complex. Although calcium clearly induces a lateral phase separation in the DMPA/DMPC-d54 system at pH 7.5 (Kouaouci, R., J.R. Silvius, I. Grah, and M. Pezolet. 1985. Biochemistry. 24:7132-7140), no distinct lateral segregation of the lipid components is observed in the mixed DMPA/DMPC-d54 lipid system in the presence of either ferricytochrome c or the sodium and calcium cations at pH 4.0. However, domain formation, consisting of regions rich in DMPA and DMPC-d54, respectively, is suggested for the calcium binary lipid mixture at pH 4.0 by the different values for Tm and AT characterizing the DMPA and DMPC-d54 species.Spectral evidence strongly suggests that ferricytochrome c also induces domain formation in the ternary lipid-protein mixtures at pH 7.0, but only for the sodium cation.

Rapid responses to stress in Eurytemora affinis

E. affinis can adjust to temperature stress in a matter of hours. Adaptation is greater in a varying temperature than in a constant temperature, consistent with the estuarine habitat of this calanoid. The species has the capacity to adjust both in the short-term as individuals and also genetically over a number of generations. The adjustments have been examined at several levels of organization. In whole copepods the time an individual becomes comatose when exposed to a 32 °C temperature and increasing by 1/2 °C at 5 min. intervals, has been used as a repeatable assay and gives a good prediction of survival at 30 °C, the ecological limit of the species in Chesapeake bay, USA. At the molecular and cellular levels, two adaptive mechanisms which have been observed in temperature stressed copepods are the synthesis of novel proteins and phase changes in plasma membrane lipids. Both of these mechanisms have potential for further understanding the adaptation of Eurytemora to variable temperatures. They may also have application as indicators of sublethal stress.

ESR of diphenylantimony radical in a triphenylantimony single crystal

Ultraviolet irradiation of a single crystal of triphenylantimony produces an antimony centered radical with an anisotropic 121Sb coupling having principal absolute values of 704, 352, and 352 MHz and a g tensor with the same principal axis as the hyperfine interaction and components of 1.998, 2.045, and 2.065. This radical has been assigned to the diphenyl antimony radical with approximately zero s and 0.7 p character. The linewidth of the spectra are consistent with unresolved hyperfine structure due to spin density delocalized on the phenyl rings.

Preparation of Carotenoid Enriched Vesicles of Monogalactosyldiacylglycerol and Digalactosyldiacylglycerol for Use in Raman Studies to Investigate Leaf Canopy Damage

Summary New methods were developed for the semi-preparative isolation of galactolipids (GL) monogalactosyl-diacylglycerol (MGDG), and digalactosyldiacylglycerol (DGDG) from Glycine max L. Merr. (soybean) leaf tissue. Model membranes composed of isolated MGDG and DGDG were enriched with the natural carotenoid, lutein and subjected to vibrational spectroscopic analysis as a function of temperature. The carotenoid was introduced to provide a resonance Raman (rR) indicator that would be sensitive to the macroscopic environment of the lipid assembly. The preliminary Infra-red (IR), Raman and rR spectra in the ordered gel phase and in the relatively disordered liquid crystalline (lc) phase of the phospholipid (PL) model membrane system distearoylphosphatidylcholine (DSPC) and spectra of the GL model membrane systems, that span the temperature range of -30 °C to 0°C, indicate that vesicles prepared by this method provide relevant model membrane systems.