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E Oldfield

Publications and source records attributed to E Oldfield.

At least 37 records · Page 2Linked to original sources
PubMed1998

A carbon-13 nuclear magnetic resonance spectroscopic study of inter-proton pair order parameters: a new approach to study order and dynamics in phospholipid membrane systems.

We report a simple new nuclear magnetic resonance (NMR) spectroscopic method to investigate order and dynamics in phospholipids in which inter-proton pair order parameters are derived by using high resolution 13C cross-polarization/magic angle spinning (CP/MAS) NMR combined with 1H dipolar echo preparation. The resulting two-dimensional NMR spectra permit determination of the motionally averaged interpair second moment for protons attached to each resolved 13C site, from which the corresponding interpair order parameters can be deducted. A spin-lock mixing pulse before cross-polarization enables the detection of spin diffusion amongst the different regions of the lipid molecules. The method was applied to a variety of model membrane systems, including 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/sterol and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/sterol model membranes. The results agree well with previous studies using specifically deuterium labeled or predeuterated phospholipid molecules. It was also found that efficient spin diffusion takes place within the phospholipid acyl chains, and between the glycerol backbone and choline headgroup of these molecules. The experiment was also applied to biosynthetically 13C-labeled ergosterol incorporated into phosphatidylcholine bilayers. These results indicate highly restricted motions of both the sterol nucleus and the aliphatic side chain, and efficient spin exchange between these structurally dissimilar regions of the sterol molecule. Finally, studies were carried out in the lamellar liquid crystalline (L alpha) and inverted hexagonal (HII) phases of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE). These results indicated that phosphatidylethanolamine lamellar phases are more ordered than the equivalent phases of phosphatidylcholines. In the HII (inverted hexagonal) phase, despite the increased translational freedom, there is highly constrained packing of the lipid molecules, particularly in the acyl chain region.

Biophysical Phenomena↗
PubMed1997

Assignment and analysis of fluorine nuclear magnetic resonance spectra of 4-fluorotryptophan myoglobins and hemoglobins.

We have obtained the 470 MHz 19F NMR spectra of wild type [4-F]Trp-labeled myoglobins (MbCO, MbO2, deoxyMb, metMb, and MbCN) and hemoglobins (HbCO, HbO2, and deoxyHb), as well as those of several mutants (W7F Mb, betaW15F Hb, betaW37S Hb, and betaY130F Hb, all as the carbonmonoxy adducts), prepared via site-directed mutagenesis. The maximum observed chemical shift range induced by folding is 6.4 ppm. Using a multipole shielding polarizability-local reaction field approach, we have computed the electrostatic field contributions to the fluorine shielding. For residues which do not have F atoms in contact with neighboring groups, we find an approximately 1 ppm mean square deviation in shift from experiment, with the R2-like structure of HbCOA being in very close accord with experiment.

Animals↗
PubMed1997

Antiproliferative effects of cyclopentenyl cytosine (NSC 375575) in human glioblastoma cells.

Cyclopentenyl cytosine (CPEC) exerts an antiproliferative effect against a wide variety of human and murine tumor lines, including a panel of human gliosarcoma and astrocytoma lines. This effect is produced primarily by the 5'-triphosphate metabolite CPEC-TP, an inhibitor of cytidine-5'-triphosphate (CTP) synthase (EC 6.3.4.2). Because previous studies with human glioma cell lines utilized cells in long-term tissue culture, we have undertaken to determine whether the activity of CPEC in such model systems is also demonstrable in freshly excised human glioblastoma cells. Glioma cells obtained at surgery and in log phase growth were exposed to the drug at levels ranging from 0.01 to 1 microM for 24 h, and CPEC-TP and CTP levels were determined by HPLC. Dose-dependent accumulation of CPEC-TP was accompanied by a concomitant decrease in CTP pools, with 50% depletion of the latter being achieved at a CPEC level of ca. 0.1 microM. Human glioma cell proliferation was inhibited 50% by 24-h exposure to 0.07 microM CPEC. Postexposure decay of CPEC-TP was slow, with a half-time of 30 h. DNA cytometry showed a dose-dependent shift in cell cycle distribution, with an accumulation of cells in S-phase. The pharmacological effects of CPEC on freshly excised glioblastoma cells are quantitatively similar to those seen in a range of established tissue culture lines, including human glioma, colon carcinoma, and MOLT-4 lymphoblasts, supporting the recommendation that the drug may be advantageous for the treatment of human glioblastoma.

Adult↗
PubMed1996

Recent progress in understanding chemical shifts.

In the past three or four years computer hardware and software developments have reached the stage where the nuclear magnetic resonance (NMR) spectra of many molecular systems can now be accurately evaluated. Detailed analysis of chemical shifts may soon become a routine part of solid (and liquid) state NMR structure prediction in chemistry and biology, and this Article covers the development of the topic from its earliest beginnings.

Carbon Isotopes↗
PubMed1995

Molecular order and dynamics of phosphatidylcholine bilayer membranes in the presence of cholesterol, ergosterol and lanosterol: a comparative study using 2H-, 13C- and 31P-NMR spectroscopy.

We report the results of a comparative study of the molecular order and dynamics of phosphatidylcholine (PC) bilayer membranes in the absence and presence of cholesterol, ergosterol and lanosterol, using deuterium (2H) nuclear magnetic resonance (NMR) of deuterated phospholipid molecules, in addition to solid state 13C and 31P-NMR. Using dimyristoylphosphatidylcholines (DMPCs) specifically labeled at positions 2', 3', 4', 6', 8', 10' and 12' of the sn-2 chain together with the perdeuterated 2-[2H27]DMPC derivative, the order profile for 9 of the 13 methylene groups of the sn-2 chain was established at 25 degrees C for DMPC, DMPC/cholesterol, DMPC/ergosterol and DMPC/lanosterol membranes, at a fixed sterol/phospholipid mol ratio of 30%, and in the presence of excess water. The overall ordering effects were found to be ergosterol > cholesterol >> lanosterol. Transverse relaxation (T2e) studies of these systems indicated that while for DMPC, DMPC/cholesterol and DMPC/ergosterol the relative relaxation rates were in qualitative agreement with models which assume cooperative motions of the bilayer molecules as the main relaxation mechanism, those in DMPC/lanosterol were anomalously high, suggesting alterations of lipid packing. Using dipalmitoylphosphatidylcholine (DPPC) deuterated at the trimethylammonium group of the choline moiety, we found that the differential ordering and motional effects induced by the sterols in the acyl chains were also reflected in the headgroup, both in the gel (L beta) and liquid-crystalline phases. 13C and 1H spin dynamics studies of these systems, including cross-polarization, rotating frame longitudinal relaxation and dipolar echo relaxation rates showed that the mobility of the different regions of the phospholipid molecules in the binary lipid systems were inversely correlated with the ordering effects induced by the sterols. A novel combination of C-D bond order parameters (obtained by 2H-NMR) and 13C-1H cross polarization rates confirmed these results. The effects of the same sterols at the same molar proportion on the unsaturated lipid 1-[2H31]palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (2H31-POPC) at 25 and 35 degrees C were different from those observed on DMPC and showed ordering effects which are largest for cholesterol, while ergosterol and lanosterol produced significantly smaller effects. Transverse relaxation studies indicate that while cholesterol does not perturb cooperative motions in POPC, both ergosterol and lanosterol do. Again, high-resolution solid state 13C-NMR studies support the conclusions of the 2H-NMR experiments.(ABSTRACT TRUNCATED AT 400 WORDS)

Carbon Isotopes↗
PubMed1995

Chemical shifts and three-dimensional protein structures.

During the past three years it has become possible to compute ab initio the 13C, 15N and 19F NMR chemical shifts of many sites in native proteins. Chemical shifts are beginning to become a useful supplement to more established methods of solution structure determination, and may find utility in solid-state analysis as well. From 13C NMR, information on phi, psi and chi torsions can be obtained, permitting both assignment verification, and structure refinement and prediction. For 15N, both torsional and hydrogen-bonding effects are important, while for 19F, chemical shifts are primarily indicators of the local charge field. Chemical shift calculations are still slow, but shielding hypersurfaces - the shift as a function of the dihedral angles that define the molecular conformation - are becoming accessible. Over the next few years, theoretical and computer hardware improvements will enable more routine use of chemical shifts in structural studies, including the study of metal-ligand interactions, the analysis of drug and substrate binding and catalysis, the study of folding/unfolding pathways, as well as the characterization of conformational substates. Rather than simply being a necessary prerequisite for multidimensional NMR, chemical shifts and chemical shift non-equivalence due to folding are now beginning to be useful for structural characterization.

Computer Simulation↗
PubMed1995

1H, 13C and 15N chemical shift referencing in biomolecular NMR.

A considerable degree of variability exists in the way that 1H, 13C and 15N chemical shifts are reported and referenced for biomolecules. In this article we explore some of the reasons for this situation and propose guidelines for future chemical shift referencing and for conversion from many common 1H, 13C and 15N chemical shift standards, now used in biomolecular NMR, to those proposed here.

Carbon Isotopes↗
PubMed1995

129Xe magic-angle spinning spectra of xenon in zeolite NaA. Direct observation of mixed clusters of co-adsorbed species.

We present the first demonstration that the individual 129Xe resonances corresponding to Xen (n = 1-8) clusters inside the alpha-cages of zeolite NaA can be narrowed under magic-angle spinning (MAS). Under these high-resolution conditions we also observe upon addition of Kr the individual peaks corresponding to mixed clusters, XenKrm, inside the alpha-cages, which will allow the first direct determination of the distribution of co-adsorbates in a microporous solid. Under MAS the chemical shifts of the Xen clusters are shown to be highly sensitive to "disorder" in the zeolite and provides new, quantitative information about the presence of alpha-cages of several types.

Magnetic Resonance Spectroscopy↗
PubMed1994

Fluorine-19 nuclear magnetic resonance spectroscopic study of fluorophenylalanine- and fluorotryptophan-labeled avian egg white lysozymes.

We report the 470-MHz (11.7 T) 19F solution nuclear magnetic resonance (NMR) spectra of 2-, 3-, and 4-fluorophenylalanine incorporated into the egg white lysozymes (EC 3.2.1.17) of chicken, pheasant, and duck, as well as spectra of 4-fluorotryptophan incorporated into chicken, California valley quail, and Bob White quail and 5- and 6-fluorotryptophan-labeled chicken lysozyme. The 19F solution NMR spectrum of [4-F]Phe hen egg white lysozyme (HEWL) consists of three sharp resonances, which span a total chemical shift range of 4.8 ppm (at p2H = 6.1). For [3-F]Phe HEWL, the chemical shift range is much smaller, 1.0 ppm (at p2H = 5.9), due presumably to the occurrence of fast phenyl ring flips about the C beta-C gamma bond axis. For [2-F]Phe HEWL, six resonances are observed, spanning a chemical shift range of 7.4 ppm (at p2H = 5.8), due to slow C beta-C gamma ring flips, i.e., both ring-flip isomers appear to be "frozen in" because of steric hindrance. Rotation of the [2-F]Phe residues remains slow up to 55 degrees C (p2H = 4.7). With the [F]Trp-labeled proteins, we find a maximal 14.6-ppm shielding range for [4-F]Trp HEWL but only a 2.8- and 2.4-ppm range for [5- and 6-F]Trp HEWL, respectively, due presumably to increased solvent exposure in the latter cases. Guanidinium chloride denaturation causes loss of essentially all chemical shift nonequivalence, as does thermal denaturation. Spectra recorded as a function of pH show relatively small chemical shift changes (< 1.4 ppm) over the pH range of approximately 1.2-7.8.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Correlation between 15N NMR chemical shifts in proteins and secondary structure.

An empirical correlation between the peptide 15N chemical shift, delta 15Ni, and the backbone torsion angles phi i, psi i-1 is reported. By using two-dimensional shielding surfaces delta (phi i, psi i-1), it is possible in many cases to make reasonably accurate predictions of 15N chemical shifts for a given structure. On average, the rms error between experiment and prediction is about 3.5 ppm. Results for threonine, valine and isoleucine are worse (approximately 4.8 ppm), due presumably to chi 1-distribution/gamma-gauche effects. The rms errors for the other amino acids are approximately 3 ppm, for a typical maximal chemical shift range of approximately 15-20 ppm. Thus, there is a significant correlation between 15N chemical shift and secondary structure.

Amino Acids↗
PubMed1994

Aluminum to oxygen cross-polarization in alpha-Al2O3 (corundum).

Experimental results concerning the cross-polarization of two quadrupolar, I = 5/2 nuclei are presented. It is shown in particular that 27Al spins in alpha-Al2O3 (corundum) can be spin-locked, and magnetization transferred to the 17O spin system. Cross-polarization between pairs of quadrupolar nuclei can cause considerable signal-to-noise ratio gains over conventional one- (or two-) pulse excitation and should be of use in investigating a number of inorganic solids.

Aluminum↗
PubMed1993

Editing 13C-NMR spectra of membranes.

We report the carbon-13 'magic-angle' sample-spinning nuclear magnetic resonance (NMR) spectra of several lipid-water systems, under a variety of radiofrequency excitation conditions. Our results show that complex lipid or membrane spectra can be greatly simplified by using 'spectral editing' techniques. For example, in a 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)-water mesophase, the glycerol (C-1, C-2 and C-3) carbons are readily distinguished from the headgroup C alpha, C beta and C gamma carbons, on the basis of their mix-time behavior in a cross-polarization (CP) experiment, while in the more complex DMPC/cholesterol-water system, many of the more rigid cholesterol carbon resonances can be edited from the phospholipid peaks. In very complex systems, such as human myelin membranes, editing permits the unambiguous observation of the mobile lipid headgroup carbon resonances, as well as the much more rigid sterol ring carbons. We also report the observation of a large differential CP due to C-H vector 'magic-angle' orientational effects in the DMPC/desipramine system. Thus, both motional or orientational reduction of the C-H dipolar interaction can lead to considerable simplifications of complex membrane spectra, and are of interest from both spectral assignment and membrane dynamics aspects.

Cell Membrane↗