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Biomedical subjects

G C Levy

Publications and source records attributed to G C Levy.

At least 37 records · Page 2Linked to original sources
PubMed1986

Phosphorus-31 nuclear magnetic resonance measurements of 2,3-DPG degradation in human adult and cord blood erythrocytes.

We have measured the percent change in 2,3-diphosphoglycerate (DPG) concentration in cord blood erythrocytes and adult erythrocytes during incubation at 37 degrees C using 31P nuclear magnetic resonance spectroscopy. DPG concentrations remained unchanged during the first 4.0 +/- 0.9 h (mean + SD) for cord blood erythrocytes and 6.0 +/- 1.0 h for adult blood erythrocytes. After this plateau phase, cord blood and adult erythrocyte DPG concentrations decreased at a rate of 11.6 +/- 0.4 and 11.4 +/- 0.5% of the baseline DPG concentration per hour, respectively. There was no statistically significant difference in the percent rate of DPG degradation when comparing cord blood erythrocytes to adult erythrocytes during either the plateau phase or the decay phase. Starting at the onset of the decay phase, the neonate 3-P DPG resonance shifted downfield at a rate of 0.03 +/- 0.006 ppm per hour; the adult 3-P DPG resonance shifted downfield at a rate of 0.02 +/- 0.003 ppm per hour.

2,3-Diphosphoglycerate↗
PubMed1985

51V-n.m.r. analysis of the binding of vanadium(V) oligoanions to sarcoplasmic reticulum.

The binding of mono- and oligo-vanadates to sarcoplasmic reticulum was analysed by 51V-n.m.r. spectroscopy. The observations indicate that, in addition to monovanadate, the di-, tetra- and deca-vanadates are also bound to sarcoplasmic-reticulum membranes with high affinity. The binding of the vanadate oligoanions may explain some of the effects of vanadates on the conformation and crystallization of Ca2+-transport ATPase.

Animals↗
PubMed1985

Conformations of bound nucleoside triphosphate effectors in aspartate transcarbamylase. Evidence for the London-Schmidt model by transferred nuclear Overhauser effects.

Transferred nuclear Overhauser effects were used to determine the conformations of ATP, CTP, and ITP bound to the regulatory site of aspartate transcarbamylase. The results are in accord with the predictions of the London-Schmidt model [London, R. E., & Schmidt, P. G. (1972) Biochemistry 11, 3136] and show that ATP and CTP bind in the anti conformation while ITP binds in the syn conformation.

Allosteric Site↗
PubMed1985

Computer systems for laboratory networks and high-performance NMR.

Modern computer technology is significantly enhancing the associated tasks of spectroscopic data acquisition and data reduction and analysis. Distributed data processing techniques, particularly laboratory computer networking, are rapidly changing the scientist's ability to optimize results from complex experiments. Optimization of nuclear magnetic resonance spectroscopy (NMR) and magnetic resonance imaging (MRI) experimental results requires use of powerful, large-memory (virtual memory preferred) computers with integrated (and supported) high-speed links to magnetic resonance instrumentation. Laboratory architectures with larger computers, in order to extend data reduction capabilities, have facilitated the transition to NMR laboratory computer networking. Examples of a polymer microstructure analysis and in vivo 31P metabolic analysis are given. This paper also discusses laboratory data processing trends anticipated over the next 5-10 years. Full networking of NMR laboratories is just now becoming a reality.

Computers↗
PubMed1984

Use of gated perfusion to study early effects of anoxia on cardiac energy metabolism: a new 31P NMR method.

A novel 31P NMR method is described that is capable of determining rapid changes in the intracellular levels of various phosphorus-containing compounds in an isolated, perfused working rat heart. This technique involves the gating of 31P NMR measurements to a heart that is alternately perfused with a modified Krebs-Henseleit medium containing 10 mM pyruvate and equilibrated with either 95% O2/5% CO2 or 95% N2/5% CO2. The experimental design allows up to three NMR measurements to be made during a single O2/N2 perfusion cycle. When these measurements are repeated at different intervals during the cycle, rapid changes in metabolite levels can be determined. Preliminary studies have shown that hearts remain hemodynamically stable to the aerobic/anoxic perfusion cycle as judged by heart rate, peak systolic pressure, aortic output, and coronary flow for at least 80 min in the magnet when subjected to cycle times of 4.5-s O2 and 1.5-s N2 perfusions. NMR measurements made under these conditions showed that a transition from full aerobic perfusion to this cycle revealed a new steady state, with an increased inorganic phosphate level from 6% total observable phosphorus to 10% and a possibly significant decreased measurement of creatine phosphate level (from 35 to 31%). Comparison of individual NMR measurements made during this perfusion cycle shows apparent rapid cyclical variations in intracellular pH and the levels of Pi, ATP, and NAD(H). These changes, expressed as variations above and below mean values measured during the cycle, showed that (a) intracellular pH, as measured by the chemical shift of Pi, reversibly decreases by more than 0.1 pH unit within 0.5-1 s following maximal anoxic perfusion and (b) coincident with a decrease in intracellular pH, Pi levels increased by a maximum of 30-40% whereas ATP levels decreased by a maximum of 15-20%. The amount of total observable phosphorous detected during the cycle is essentially constant. Unexpectedly, creatine phosphate levels are most stable, indicating that their levels are being maintained at the expense of ATP. Also unexpected is the finding that NAD(H) levels varied from maximal to undetectable levels during the perfusion cycle. The current method of aerobic/anoxic perfusion is capable of resolving metabolic events much faster than previous NMR methods and yielding information that is unobtainable by any other technique.

Adenosine Triphosphate↗
PubMed1984

31P-NMR analysis of the B to Z transition in double-stranded (dC-dG)3 and (dC-dG)4 in high salt solution.

In 4M NaCl solutions (dC-dG)n (n = 3,4; approximately 9 mM) exist as a mixture o +/- B and Z forms. The low and high field components of two 31P NMR resonances originating from internal phosphodiester groups are assigned to the GpC and CpG linkages, respectively. Low temperatures stabilize the Z-forms, which completely disappear above 50 degrees C (n = 3) and 65 degrees C (n = 4). delta H = -44 and -17 kJ/mol for B to Z transition in the hexamer and octamer duplexes, respectively. Temperature dependent changes (0-50 degrees C range) in the spin-lattice relaxation times at 145.7 MHz are distinctly different for the 31P nuclei o +/- GpC and CpG groups. The relaxation data can be explained by assuming that the GpC phosphodiester groups undergo more local internal motion than do the CpG groups.

Calorimetry↗
PubMed1984

Conformation and dynamics of short DNA duplexes: (dC-dG)3 and (dC-dG)4.

Natural abundance 13C NMR spectra of duplexed (dC-dG)3 and (dC-dG)4 exhibit resolved resonances for most of the carbons at 0.1M NaCl in aqueous solution. Large transitions in chemical shift for many of the hexamer carbons (up to 1.8 ppm) are observed in variable temperature measurements. Determination of spin-lattice relaxation times and nuclear Overhauser enhancements in 0.1M NaCl indicate that the duplexes tumble almost isotropically, with overall correlation times near 5 nsec; the sugar carbons experience more rapid local motions than do the base carbons. The relaxation data are also consistent with the most rapid local motions occurring at the chain-terminal residues, especially in the Cyd(1) sugar. 4M NaCl causes changes in the 13C chemical shifts of most of the guanine base carbons, and rearrangements in the deoxyribose carbon shifts; this is consistent with changes predicted by a salt-induced B to Z transition, viz. conversion of the guanylates from the anti to syn range about the glycosyl bond, and from the S to N pseudorotational state of the deoxyribose ring.

DNA↗
PubMed1983

Conformations of nicotinamide coenzymes bound to dehydrogenases determined by transferred nuclear Overhauser effects.

Transferred nuclear Overhauser enhancement was used to examine the conformation of NAD+ and NADP+ bound to glucose-6-phosphate dehydrogenase and glutamate dehydrogenase and of NAD+ bound to lactate dehydrogenase. The results demonstrate that the conformation of the nicotinamide-ribose bond is anti for dehydrogenases with A stereospecificity and syn for dehydrogenases with B stereospecificity. In those dehydrogenases that bind both NAD+ and NADP+, significant differences occur in the conformations of the bound nicotinamide coenzymes.

Animals↗
PubMed1983

Spontaneous ordering of DNA. Effects of intermolecular interactions on DNA motional dynamics monitored by 13C and 31P nuclear magnetic resonance spectroscopy.

Effects of intermolecular DNA interactions on the motional dynamics of defined length (147-, 234-, and 437-nucleotide pair (np)) double-stranded DNA were examined by 13C and 31P nuclear magnetic resonance spectroscopy. At a critical high concentration varying inversely with length, DNA undergoes a spontaneous transition to an ordered, liquid crystalline-like state. Ordering is accompanied by the appearance of distinct opalescence, and an increase in solution viscosity. An apparent standard heat of fusion of -38 kcal/mol of helix (-0.13 kcal/mol of DNA phosphate) and an entropy change of -0.13 entropy units (per mol of helix) were determined from the temperature dependence of the phase transition of 147-np DNA. The average phosphodiester configuration, monitored by Raman spectroscopy, was typical of B-form DNA above and below the phase transition. NMR spectra and relaxation data show that intermolecular interactions are strong at concentrations well below the phase transition and cause stepwise uncoupling of internal motions at specific sites. Motions of the exocyclic C5' carbon, but not other sugar carbons, are frozen in approximately 50% of 147-np DNA molecules at concentrations as low as 6.5 mg/ml. Motions of backbone ring carbons (C3', C4') are frozen in a progressively larger fraction of molecules at concentrations above 46 mg/ml. Rapid C2' motions are unaffected below the critical concentration (193 mg/ml at 32 degrees C), and still occur in the ordered phase. We conclude that rapid internal motions of DNA monitored by NMR consist mostly of coupled, periodic bending deformations and partially uncoupled local motions within the sugar ring. Rapid, extensive C2' motions can occur without strong coupling to other sugar carbon motions. From a comparison of 13C and 31P NMR data, we conclude that 31P NMR at best yields an incomplete representation of DNA dynamics.

Animals↗
PubMed1983

Understanding DNA conformational dynamics: answering questions and questioning answers.

Phosphorus-31 and especially Carbon-13 NMR measurements have recently become primary input to the understanding of DNA solution dynamics. While the 31P measurements are inherently easier, the quality of 31P dynamics information is suspect and therefore 13C measurements are preferred. In fact, it is necessary to obtain several kinds of 13C data (T1s, NOE's, linewidths, integrated peak intensities) over a wide range of magnetic fields (13C NMR frequencies) in order to identify major features of DNA internal motions. Further information comes from variation of temperature and DNA fragment length and/or concentration. Most of our 13C measurements have been performed at 37.7-90.6 MHz on fully double stranded monomer size (147 base pair) DNA at concentrations in phosphate buffer of approximately less than 10 to approximately greater than 200 mg ml-1; temperatures studied range from 6 to 55 degrees C. Other measurements have been performed on monomer-size single-stranded DNA at 85 and 92 degrees. The large data set we have acquired appears to answer some important questions about the nature and extent of DNA overall and internal motional dynamics. However, the picture remains incomplete and a number of questions arise from these results: 1. Overall motion of the double stranded DNA fragments follows expected hydrodynamic behavior; 2. Restricted but rapid internal motion along the DNA structure is well represented by a spaghetti-like wobbling-in-a-cone model; 3. DNA-DNA Interactions and solvent ordering, present at relatively low DNA concentrations, partially quench the internal motion, consistent with hinge-model structural changes (and the spaghetti model above) but not as compatible with in-plane torsional motion models; 4. The deoxyribose C-2' sites undergo additional motion which is partially uncoupled from the internal wobbling motions: 5. At high DNA concentrations, a phase transition occurs, resulting in ordered structures which drastically affect DNA internal dynamics; 6. DNA interacting with ethidium does not greatly change its conformational mobility; 7. DNA interacting with Hg2+ ions shows less than anticipated change in internal DNA dynamics. The remaining challenge is to interpret our current results in terms of specific conformational processes and to understand why the conformational mobility of double stranded DNA is relatively unhindered by major structural perturbants such as intercalating ethidium and mercury ion.

DNA↗
PubMed1982

13C NMR relaxation and conformational flexibility of the deoxyribose ring.

13C T1's and NOE's have been measured for all protonated carbons of 2'-deoxy-D-ribose (2'-d-ribose), 2'-deoxyadenosine-5'-monophosphate (5'-dAMP), thymidine-3'-monophosphate (3'-TMP) and thymidine-5'-monophosphate (5'-TMP) in D2O solutions. In all of the deoxy sugars examined, NT1 values for C-2' are significantly larger than the values for the remaining carbons. This result is interpreted in terms of rapid puckering motion of C-2'. By contrast, NT1 values measured in ribose are found to be equal, within experimental error. The results are compared with analogous data obtained for the five membered pyrrolidine ring of proline and with results for DNA itself.

DNA↗
PubMed1981

Recent developments in nuclear magnetic resonance spectroscopy.

Nuclear magnetic resonance spectroscopy is a powerful and versatile technique that yields information related to molecular structure, interactions, and dynamics. Methods are currently being developed for real-time monitoring of metabolic processes in vivo and for noninvasive detection of disease and abnormality in living animals. Other nuclear magnetic resonance techniques are providing entirely new approaches for analyses of complex chemical systems. The increased power and popularity of nuclear magnetic resonance spectroscopy today are due to many developments in instrumentation and methods that have occurred over the 35-year history of the technique. The most important single advance, particularly in recent years, has been increased sensitivity. Concurrent improvements in effective spectral resolving power and an array of new methods and applications have also contributed to elevating nuclear magnetic resonance spectroscopy to its present position as one of the premier analytical techniques.

Fourier Analysis↗
PubMed1981

Carbon 13 spin-lattice relaxation, linewidth, and nuclear Overhauser enhancement measurements of nucleosome length DNA.

Carbon 13 NMR data were obtained at four magnetic fields for double-stranded DNA samples 120 and 160 nucleotide pairs long. Spectral linewidths are several-fold smaller than predicted for overall rigid rod rotation and increase with increasing field, indicating significant chemical shift dispersion contributions to high field linewidths. Spin-lattice relaxation times are short (e.g. T1 values for CH carbon atoms are approximately 0.5 s at 67.9 MHz), and increase with increasing field. Nuclear Overhauser effects of 0.6 to 0.8 (theoretical maximum = 2.0) were measured for the same protonated carbon atoms at 100.6 MHz. These short T1 values nd relatively large nuclear Overhauser effects show that double-stranded DNA undergoes rapid internal motions with effective correlation times of a few nanoseconds. Preliminary data indicate that DNA base carbon motions occur on essentially the same time scale as sugar motions, thus arguing against independent flexibility of the DNA backbone. Linewidths decreased 2- to 5-fold and nuclear Overhauser effects doubled, upon heat denaturation of DNA, as expected for increased motion. On the other hand, NT1 values of native and denatured DNA were nearly identical, suggesting insensitivity of spin-lattice relaxation times to motions in the nanosecond range in these relatively stiff chains.

Animals↗
PubMed1977

The influence of glycyl residues on the flexibility of peptide hormones in solution. A 13C-nuclear-magnetic-resonance study of luteinizing hormone-releasing hormone (luliberin) and its des-glycinamide10 N-ethylamide analog.

13C nuclear magnetic resonance spectroscopy in used to gain information on the flexibility of the backbone in peptide hormones and peptide hormone analogs. 13C spin-lattice relaxation times (T1) were measured on luliberin, the luteinizing-hormone-releasing hormone and des(Gly-NH2)10-luliberin-N-ethylamide in aqueous solution at 25.2 and 67.9 MHz at temperatures of 32 degrees, 40 degrees and 55 degrees C. The 13C spin-lattice relaxation times indicate increased flexibility of the peptide backbone in the immediate environment of glycyl residues in luliberin (less than Glu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2) and the hormone analog des(Gly-NH2)10-luliberin-N-ethylamide (less than Glu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-NH-CH2CH3) in aqueous solutions. 13 C NMR spectroscopy is shown to be a sensitive technique for monitoring the time-averaged conformational flexibility of peptides in solution. Activation energies (Ea) of about 25 kJ/mol were obtained for rotational reorientation of non-terminal alpha-carbons in the peptide backbone. Rotation of methyl groups was characterized by an Ea of 9.6 kJ/mol whereas reorientation of the N-terminal pyroglutamyl residue showed an Ea value of 14.6 kJ/mol. The Ea values of individual carbons in the side-chains of prolyl, arginyl and leucyl residues in the peptides were similar to those obtained for the alpha-carbon of the same amino acid residue in the peptide backbone of the hormones.

Calorimetry↗
PubMed1976

A carbon-13 nuclear magnetic resonance study of hyperactive and hypoactive derivatives of luteinizing hormone-releasing hormone: des-Gly-NH210-[DLeu6]LH-RH ethylamide and des-Gly-NH210-[L-Leu6]LH-RH ethylamide.

The carbon-13 (13C) nuclear magnetic resonance (NMR) behavior of the hyperactive analogue of luteinizing-hormone-releasing hormone (LH-RH), des-Gly-NH2(10)-[D-Leu6]LH-RH ethylamide, has been studied. The spectra are compared with those of the hypoactive des-Gly-NH2(10)-[L-Leu6]LH-RH ethylamide. Spin-lattice relaxation times (T1) of 13C at natural abundance have been obtained for both peptides at 25 and 68 MHz in aqueous solution and used as monitors of flexibility at each carbon atom in the peptide backbone and side chains. The [D-Leu6]-and [L-Leu6]-sub-stituted analogues show equal degrees of freedom within the peptide backbone and side chains. Substitution of the glycine amide moiety in position 10 by an ethylamide group causes an increase in flexibility of the backbone at that position. The increase in flexibility is not transmitted to the adjacent prolyl residue. The C-terminal peptide link at position 10 is proposed to restrict the flexibility of the prolyl residue in position 9, thus maintaining the motional characteristics of the LH-RH peptide backbone in the des-Gly-NH2(10)-[Leu6]LH-RH ethylamide analogues. There were no significant differences in the 13C chemical shifts or T1 values between the hypoactive and hyperactive analogues in aqueous solution which would explain the differences in biological activity. Interaction between receptor and hormone may thus be required to induce, or select, the biologically active conformation of the hormone.

Amides↗