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Biomedical subjects

D H Turner

Publications and source records attributed to D H Turner.

At least 109 records · Page 6Linked to original sources
PubMed1985

Fluorescence-detected circular dichroism of ethidium in vivo and bound to deoxyribonucleic acid in vitro.

Fluorescence-detected circular dichroism (FDCD) spectra are reported for ethidium in Escherichia coli cells and bound to E. coli DNA in vitro. FDCD bands are observed at 325 and 385 nm. These bands change amplitude as the ethidium to DNA ratio changes. Spectra are similar for in vivo and in vitro measurements. However, the bands at 325 and 385 nm disappear when ethidium binds to macromolecules without intercalating between base pairs. The results demonstrate that FDCD spectra can be measured in cell suspensions and indicate that ethidium binds to nucleic acids in E. coli cells by intercalation.

Chemical Phenomena↗
PubMed1985

Solvent effects on the stability of A7U7p.

The thermodynamics of double-helix formation were measured spectrophotometrically for A7U7 in water at 1 M NaCl and for A7U7p in a variety of solvent mixtures and salt. Comparison of the A7U7 results with calorimetric measurements indicates duplex formation involves intermediate states. For A7U7p between 0.06 and 0.55 M Na+, dTm/d(log [Na+]) = 17.4 degrees C, similar to the value of 19.6 degrees C for poly-(A).poly(U) [Krakauer, H., & Sturtevant, J. M. (1968) Biopolymers 6, 491-512]. At 1 M NaCl, the A7U7p duplex is most stable in 100% water. For 10 mol % solutions, the order for A7U7p duplex stability is ethylene glycol greater than glycerol greater than ethanol greater than 2-propanol greater than dimethyl sulfoxide greater than 1-propanol greater than formamide greater than N,N-dimethylformamide greater than urea greater than dioxane. Comparison of changes in stability and thermodynamic parameters with literature results for proteins suggests proteins and A7U7p interact differently with solvent. The results suggest hydrophobic bonding is not a major contributor to the stability of the A7U7p duplex. Comparisons with bulk solvent surface tension suggest the energy of cavity formation is also not a major contributor to duplex stability.

Calorimetry↗
PubMed1985

Substituted benzamides with conformationally restricted side chains. 1. Quinolizidine derivatives as selective gastric prokinetic agents.

The gastric prokinetic action of metoclopramide may not be primarily due to its dopamine antagonist activity. The present aim was to obtain a selective gastric prokinetic agent lacking dopamine antagonist activity by conformationally restricting the side chain of metoclopramide. In a series of quinolizidinylbenzamides, only compounds with the benzamide moiety at position 2 of the quinolizidine ring retain gastric activity. Of these 2-substituted compounds, the 2 alpha, 9a alpha isomer has potent selective gastric prokinetic activity with only weak dopamine antagonist properties. Spectroscopic data show that the quinolizidine ring preferentially adopts a trans chair-chair conformation with an axial benzamide moiety. However, energy calculations indicate that, at nondopaminergic receptors controlling gastric motility, an alternative cis chair-chair conformation with an equatorial benzamide moiety cannot be ruled out.

Animals↗
PubMed1984

Thermodynamic studies of RNA stability.

Enthalpies and entropies of helix stabilization due to addition of 3' terminal unpaired nucleotides to a CCGG or GGCC core double helix are derived from UV melting studies. The results suggest stacking provides a significant fraction of the free energy of a terminal base pair. The effects of temperature, aggregation, and ionic strength on the determination of thermodynamic parameters are considered. Helix propagation parameters are revised and extended based on recent additions to the data set.

Base Composition↗
PubMed1983

Base-stacking and base-pairing contributions to helix stability: thermodynamics of double-helix formation with CCGG, CCGGp, CCGGAp, ACCGGp, CCGGUp, and ACCGGUp.

The thermodynamics of double-helix formation in 1 M NaCl have been measured spectrophotometrically for CCGG, CCGGp, CCGGAp, ACCGGp, CCGGUp, and ACCGGUp. The results indicate additional double-helical stability is conferred by the terminal unpaired bases. The 3' A stabilizes the double helix more than the 5' A or the 3' U. The increased stability is due to a more favorable enthalpy change for double-helix formation. Comparison of the thermodynamics for CCGG, ACCGGp, CCGGUp, and ACCGGUp indicates stacking interactions are somewhat more important than pairing interactions in determining the stability of the terminal AU base pairs in ACCGGUp.

Base Composition↗
PubMed1983

Nuclear overhauser studies of CCGGAp, ACCGGp, and ACCGGUp.

Nuclear Overhauser effect (NOE) measurements are reported for the nonexchangeable base and ribose 1' proton resonances of CCGGAp, ACCGGp, and ACCGGUp. The results permit assignment of these resonances to particular nucleotides in the sequences. The NOE data rule out conformations containing syn glycosidic linkages and conformations in which the purine 8 and pyrimidine 6 protons are closer to the 3' nearest-neighbor ribose 1' proton than to the 5' nearest-neighbor ribose 1' proton.

Base Sequence↗
PubMed1983

Proton magnetic resonance melting studies of CCGGp, CCGGAp, ACCGGp, CCGGUp, and ACCGGUp.

Proton magnetic resonance melting curves are reported for the base and ribose 1' nonexchangeable protons of CCGGp, CCGGAp, ACCGGp, CCGGUp, and ACCGGUp. In general, the double helices melt in a cooperative two-state transition. For CCGGp, CCGGAp, CCGGUp, and ACCGGUp, the 3' ribose exhibits conformational flexibility at lower temperatures than the other ribose residues. For ACCGGp, the 5'-C ribose is the first to exhibit flexibility. The conformations of the 3' terminal bases in CCGGAp and CCGGUp change cooperatively with the double- to single-strand transition, whereas the 5' A of ACCGGp appears to have more conformational freedom.

Base Sequence↗
PubMed1982

Laser crosslinking of E. coli RNA polymerase and T7 DNA.

The first photochemical crosslinking of a protein to a nucleic acid using laser excitation is reported. A single, 120 mJ, 20 ns pulse at 248 nm crosslinks about 10% of bound E. coli RNA polymerase to T7 DNA under the conditions studied. The crosslinking yield depends on mercaptoethanol concentration, and is a linear function of laser intensity. The protein subunits crosslinked to DNA are beta, beta' and sigma.

DNA, Viral↗
PubMed1981

Thermodynamics of (dG--dC)3 double-helix formation in water and deuterium oxide.

The thermodynamics of double-helix formation by (dG-dC)3 in H2O and D2O were measured spectrophotometrically and calorimetrically. Spectrophotometric values based on plots of inverse melting temperature vs. log of concentration are in good agreement with calorimetry. However, spectrophotometric values based on the shape of absorbance vs. temperature curves are in good agreement with calorimetry only if linear pre- and posttransition base lines are subtracted before analysis. The averages of the enthalpies measured by the three most reliable spectroscopic methods are -56.9 and -62.7 kcal/mol of helix in H2O and D2O, respectively. The corresponding calorimetric values are -59.6 and -65.8 kcal/mol of helix.

Calorimetry, Differential Scanning↗
PubMed1981

Solvent effects on thermodynamics of double-helix formation in (dG-dC)3.

The thermodynamics of double-helix formation by (dG-dC)3 have been measured in aqueous solvent mixtures containing 10 mol % methanol, ethanol, 1-propanol, formamide, N,N-dimethylformamide, or urea and 20 mol % ethanol. Optical activity measurements indicate the conformation of the double helix at 3 degree C is the same in all the solvent mixtures except 20 mol % ethanol. All the cosolvents destabilize the helix relative to water. With 10 mol % alcohol cosolvents, this destabilization is associated with a more unfavorable entropy change averaging approximately 8% and a more favorable enthalpy change averaging approximately 5%. This is consistent with a small contribution of hydrophobic bonding to stability. In contrast, the destabilization by formamide, N,N-dimethylformamide, and urea is associated with a more unfavorable enthalpy change averaging approximately 23% and a more favorable entropy change averaging approximately 21%. Since all three of these cosolvents have dipole moments larger than water, this is consistent with increased competition for dipolar interactions between the nucleic acid bases. None of the results correlate with any one bulk solvent parameter such as surface tension, viscosity, or dielectric constant. With 20 mol % ethanol, optical activity measurements are consistent with a partial B to C form transition. This is associated with a 27% less favorable enthalpy and 25% more favorable entropy for helix formation relative to water. Since the B to C transition is associated with helix dehydration, this may imply a significant contribution of bound water to stability.

Ethanol↗
PubMed1981

Solvent effects on the kinetics and thermodynamics of stacking in poly(cytidylic acid).

The Raman laser temperature-jump technique has been used to measure the kinetics of the coil to helix reaction of poly(cytidylic acid) [poly(C)] in aqueous cosolvent mixtures. The rate of helix formation has a low activation energy and is proportional to reciprocal solvent viscosity. The observations suggest helix formation is rotationally diffusion controlled. The rate of coil formation in poly(C) has an activation energy of approximately 11 kcal/mol, presumably reflecting the electronic stacking interactions which stabilize the helix. Viscous cosolvents, glycerol or sucrose, slow down the rate of coil formation; acetonitrile and formamide at 5 mol % increase the rate relative to that in water. The polar cosolvents may specifically attack a cytosine stack. The absorbance vs. temperature profiles for poly(C) are analyzed with the one-dimensional Ising model. When only optical data are used, the cooperativity parameter, sigma, and the enthalpy, delta H, cannot be uniquely determined. A method is proposed that allows determination of sigma by combining spectroscopic and calorimetric data. The values of sigma derived for poly(C) are between 0.8 and 1.0, and delta H is about -9 kcal/mol of stack. An alternative method using integration of the excess heat capacity curve and extrapolation to fully stacked and random coil species yields a delta H of -7 kcal/mol of stack.

Acetonitriles↗
PubMed1980

Study of the kinetic and structural properties of a monoclonal immunoglobulin G cryoglobulin.

The precipitation of a monoclonal IgG2 crystalline cryoglobulin (WEB) is shown to occur via a nucleation mechanism. The kinetics of precipitation fits the empirical equation 1n(t2/t1) = nù(c1/c2), where t is the time to reach half-maximum turbidity for concentration c. The effect of mild reduction of interchain disulfides on the kinetics and extent of precipitation have also been determined. Cleavage of only one or two inter heavy-heavy chain disulfide bonds appears to abolish cryoprecipitation. The extent of reduction at high reductant levels suggests that there is an extra inter heavy-heavy chain disulfide in IgG-WEB. Laser Raman spectroscopy detects no structural change between reduced (noncryoprecipitable) and native (cryoprecipitable) WEB, suggesting that perturbation of a local site is responsible for loss of cryoprecipitation.

Chemical Precipitation↗
PubMed1980

Laser temperature jump study of solvent effects of poly(adenylic acid) stacking.

The dynamics of the single-stranded, helix--coil transition of poly(adenylic acid) have been investigated by using the Raman laser temperature jump technique. The driving forces of this conformational transition have been probed by varying the cosolvent in mixed aqueous solutions. The rate of helix formation correlates well with the inverse of the solution viscosity. This correlation and the low activation barrier of approximately 4 kcal/mol for this process indicate a rotational diffusion controlled reaction. The rate of coil formation has a much higher activation barrier, presumably due to the strength of base stacking in the helix. This unstacking rate is virtually unaffected by alcohol cosolvents. A significant increase in this rate occurs when polar cosolvents such as urea, formamide, or acetonitrile are added. Absorbance changes with temperature suggest that urea specifically solvates the adenine base. The polar cosolvents appear to break up the stacked, helical conformation by interacting with the bases.

Cacodylic Acid↗
PubMed1979

Spectroscopic and kinetic analysis of a monoclonal IgG cryoglobulin. Effect of mild reduction on cryoprecipitation.

The precipitation of a monoclonal IgG2 crystalline cryoglobulin (WEB) is shown to be highly dependent on temperature and concentration. Below a critical concentration of 0.6 mg/mL there is no cryoprecipitation. The kinetics of the aggregation exhibits a concentration-dependent lag time. This evidence suggests that a nucleation event is important in the precipitation. Circular dichroism (CD) was used to investigate the conformational properties of the protein. At a low concentration (0.12 or 0.15 mg/mL), no detectable spectral changes in the far- and near-UV range were noted between 40 and 3 degrees C. However, at higher concentrations (1.21 mg/mL), a small and rapid CD change was observed in the 250-280-nm region at 3 degrees C. This indicates an intermolecular interaction that precedes the precipitation. Cryoprecipitation of WEB was also shown to be dependent on maintenance of intact interchain disulfide bonds. Only one or two interchain disulfides need be cleaved to abolish cryocrystallization and to significantly diminish the CD change at 3 degrees C. The evidence is consistent with the formation of an initial intermediate that involves interactions near the disulfide bonds in the hinge region of the cryoimmunoglobulin. In this model, cleavage of these disulfides prevents this interaction and abolishes cryoprecipitation.

Circular Dichroism↗