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M Eisenberg

Publications and source records attributed to M Eisenberg.

At least 19 recordsLinked to original sources
PubMed1992

The three-dimensional structure in solution (pH 5.8) of a DNA 9-mer duplex containing 1,N2-propanodeoxyguanosine opposite deoxyadenosine. Restrained molecular dynamics and NOE-based refinement calculations.

The solution structure of the complementary d(C1-A2-T3-G4-X5-G6-T7-A8-C9).d(G10-T11-A12-C13-A14-C15-A 16-T17-G18) nonanucleotide duplex (designated X.A 9-mer) that contains a 1,N2-propanodeoxyguanosine exocyclic adduct, X5, opposite deoxyadenosine A14 in the center of the helix at pH 5.8 is investigated by use of restrained molecular dynamics followed by NOE-based back-calculation refinement. The molecular dynamics calculation is based on 91 interresidue and 97 intraresidue interproton distance restraints derived from two-dimensional nuclear Overhauser enhancement data on the X.A 9-mer at mixing times of 50 and 250 ms [Kouchakdjian, M., Marinelli, E., Gao, X., Johnson, F., Grollman, A., & Patel, D.J. (1989) Biochemistry 28, 5647-5657]. Separate runs start from classical A and B DNA and converge to essentially identical structures (atomic root mean square difference of 0.69 A). Both structures are B-type DNA in character and satisfy the experimental distance restraints with the rms difference of only 0.001 A between the calculated and experimental interproton distances. The dynamics behavior of the A----B DNA transition is monitored and analyzed. Our results clearly indicate that the driving force of the convergence is the experimental interproton distance restraints. The molecular dynamics structures are further refined by a back-calculation dynamics which directly minimizes the difference between the observed 2D NOE intensities and those calculated by the full relaxation matrix approach. The fit of the refined structures to the NOE intensities is measured by the NOE R value, which is analogous to the crystallographic residual index. These R values of the final structures are only 0.17. The refined structures are generally B type, and their convergency improves slightly to an atomic root mean square difference of 0.64 A, despite relatively large structural shifts (approximately 1 A) which occur during the back-calculation refinement in both cases. These results suggest that the converged refined structures represent reasonable approximations of the solution structure.

Base Sequence↗
PubMed1991

Structural features of an exocyclic adduct positioned opposite an abasic site in a DNA duplex.

Structural studies have been extended to dual lesions where an exocyclic adduct is positioned opposite an abasic site in the center of a DNA oligomer duplex. NMR and energy minimization studies were performed on the 1,N2-propanodeoxyguanosine exocyclic adduct (X) positioned opposite a tetrahydrofuran abasic site (F) with the dual lesions located in the center of the (C1-A2-T3-G4-X5-G6-T7-A8-C9).(G10-T11-A12-C-13-F14-C15 -A16-T17-G-18) X.F 9-mer duplex. Two-dimensional NMR experiments establish that the X.F 9-mer helix is right-handed with Watson-Crick A.T and G.C base pairing on either side of the lesion site. NOEs are detected from the methylene protons of the exocyclic ring of X5 to the imino protons of G4.C15 and G6.C13 which flank the lesion site, as well as to the H1' and H1" protons of the cross strand F14 tetrahydrofuran moiety. These NMR results establish that the exocyclic adduct X5 is positioned between flanking G4.C15 and G6.C13 base pairs and directed toward the abasic lesion F14 on the partner strand. These studies establish that the exocyclic ring of the 1,N2-propanodeoxyguanosine adduct fits into the cavity generated by the abasic site.

Base Composition↗
PubMed1991

NMR studies of the exocyclic 1,N6-ethenodeoxyadenosine adduct (epsilon dA) opposite thymidine in a DNA duplex. Nonplanar alignment of epsilon dA(anti) and dT(anti) at the lesion site.

Two-dimensional proton NMR studies are reported on the complementary d(C-A-T-G-T-G-T-A-C).d(G-T-A-C-epsilon A-C-A-T-G) nonanucleotide duplex (designated epsilon dA.dT 9-mer duplex) containing 1,N6-ethenodeoxyadenosine (epsilon dA), a carcinogen-DNA adduct, positioned opposite thymidine in the center of the helix. Our NMR studies have focused on the conformation of the epsilon dA.dT 9-mer duplex at neutral pH with emphasis on defining the alignment at the dT5.epsilon dA14 lesion site. The through-space NOE distance connectivities establish that both dT5 and epsilon dA14 adopt anti glycosidic torsion angles, are directed into the interior of the helix, and stack with flanking Watson-Crick dG4.dC15 and dG6.dC13 pairs. Furthermore, the d(G4-T5-G6).d(C13-epsilon A14-C15) trinucleotide segment centered about the dT5.epsilon dA14 lesion site adopts a right-handed helical conformation in solution. Energy minimization computations were undertaken starting from six different alignments of dT5(anti) and epsilon dA14(anti) at the lesion site and were guided by distance constraints defined by lower and upper bounds estimated from NOESY data sets on the epsilon dA.dT 9-mer duplex. Two families of energy-minimized structures were identified with the dT5 displaced toward either the flanking dG4.dC15 or the dG6.dC13 base pair. These structures can be differentiated on the basis of the observed NOEs from the imino proton of dT5 to the imino proton of dG4 but not dG6 and to the amino protons of dC15 but not dC13 that were not included in the constraints data set used in energy minimization. Our NMR data are consistent with a nonplanar alignment of epsilon dA14(anti) and dT5(anti) with dT5 displaced toward the flanking dG4.dC15 base pair within the d(G4-T5-G6).d(C13-epsilon A14-C15) segment of the epsilon dA.dT 9-mer duplex.

Adenosine↗
PubMed1991

NMR structural studies of the ionizing radiation adduct 7-hydro-8-oxodeoxyguanosine (8-oxo-7H-dG) opposite deoxyadenosine in a DNA duplex. 8-Oxo-7H-dG(syn).dA(anti) alignment at lesion site.

Proton NMR studies are reported on the complementary d(C1-C2-A3-C4-T5-A6-oxo-G7-T8-C9-A10-C11-C12).d(G13-G14-T15- G16-A17-A18-T19- A20-G21-T22-G23-G24) dodecanucleotide duplex (designated 8-oxo-7H-dG.dA 12-mer), which contains a centrally located 7-hydro-8-oxodeoxyguanosine (8-oxo-7H-dG) residue, a group commonly found in DNA that has been exposed to ionizing radiation or oxidizing free radicals. From the NMR spectra it can be deduced that this moiety exists as two tautomers, or gives rise to two DNA conformations, that are in equilibrium and that exchange slowly. The present study focuses on the major component of the equilibrium that originates in the 6,8-dioxo tautomer of 8-oxo-7H-dG. We have assigned the exchangeable NH1, NH7, and NH2-2 base protons located on the Watson-Crick and Hoogsteen edges of 8-oxo-7H-dG7 in the 8-oxo-7H-dG.dA 12-mer duplex, using an analysis of one- and two-dimensional nuclear Overhauser enhancement (NOE) data in H2O solution. The observed NOEs derived from the NH7 proton of 8-oxo-7H-dG7 to the H2 and NH2-6 protons of dA18 establish an 8-oxo-7H-dG7(syn).dA 18(anti) alignment at the lesion site in the 8-oxo-7H-dG.dA 12-mer duplex in solution. This alignment, which places the 8-oxo group in the minor groove, was further characterized by an analysis of the NOESY spectrum of the 8-oxo-7H-dG.dA 12-mer duplex in D2O solution. We were able to detect a set of intra- and interstrand NOEs between protons (exchangeable and nonexchangeable) on adjacent residues in the d(A6-oxo-G7-T8).d(A17-A18-T19) trinucleotide segment centered about the lesion site that establishes stacking of the oxo-dG7(syn).dA(anti) pair between stable Watson-Crick dA6.dT19 and dT8.dA17 base pairs with minimal perturbation of the helix. Thus, both strands of the 8-oxo-7H-dG.dA 12-mer duplex adopt right-handed conformations at and adjacent to the lesion site, the unmodified bases adopt anti glycosidic torsion angles, and the bases are stacked into the helix. The energy-minimized conformation of the central d(A6-oxo-G7-T8).d(A17-A18-T19) segment requires that the 8-oxo-7H-dG7(syn).dA18(anti) alignment be stabilized by two hydrogen bonds from NH7 and O6 of 8-oxo-7H-dG7(syn) to N1 and NH2-6 of dA18(anti), respectively, at the lesion site.(ABSTRACT TRUNCATED AT 400 WORDS)

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1990

NMR studies of an exocyclic 1,N2-propanodeoxyguanosine adduct (X) located opposite deoxyadenosine (A) in DNA duplexes at basic pH: simultaneous partial intercalation of X and A between stacked bases.

The NMR parameters for the 1,N2-propanodeoxyguanosine (X) opposite deoxyadenosine positioned in the center of the complementary d(C1-A2-T3-G4-X5-G6-T7-A8-C9).d(G10-T11-A12-C13-A14-C15-A 16-T17-G18) X.A 9-mer duplex are pH dependent. A previous paper established protonated X5(syn).A14(anti) pairing in the X.A 9-mer duplex at pH 5.8 [Kouchakdjian, M., Marinelli, E., Gao, X., Johnson, F., Grollman, A., & Patel, D. J. (1989) Biochemistry 28, 5647-5657]; this paper focuses on the pairing alignment at the lesion site at pH 8.9. The observed NOEs between specific exocyclic CH2 protons and both the imino proton of G6 and the sugar H1' protons of C13 and A14 establish that X5 is positioned toward the G6.C13 base pair with the exocyclic ring directed between C13 and A14 on the partner strand. The observed NOE between the H2 proton of A14 and the imino proton of G4, but not G6, establishes that A14 at the lesion site is directed toward the G4.C15 base pair. NOEs are detected between all exocyclic CH2 protons of X5 and the H2 proton of A14, confirming that both X5 and A14 are directed toward the interior of the helix. The X5(anti).A14(anti) alignment at pH 8.9 is accommodated within the helix with retention of Watson-Crick pairing at flanking G4.C15 and G6.C13 base pairs. The energy-minimized conformation of the (G4-X5-G6).(C13-A14-C15) segment at pH 8.9 establishes that X5 and A14 are directed into the helix, partially stack on each other, and are not stabilized by intermolecular hydrogen bonds. The X5 base is partially intercalated between C13 and A14 on the unmodified strand, while A14 is partially intercalated between G4 and X5 on the modified strand. This results in a larger separation between the G4.C15 and G6.C13 base pairs flanking the lesion site in the basic pH conformation of the X.A 9-mer duplex. The midpoint of the transition between the protonated X5(syn).A14(anti) and X5(anti).A14(anti) conformations occurs at pH 7.6, establishing an unusually high pKa for protonation of the A14 ring opposite the X5 exocyclic adduct site. Thus, the interplay between hydrophobic and hydrogen-bonding contributions modulated by pH defines the alignment of 1,N2-propanodeoxyguanosine opposite deoxyadenosine in the interior of DNA helices.

Base Composition↗
PubMed1990

Molecular cloning of the UMP synthase gene rudimentary-like from Drosophila melanogaster.

The rudimentary-like locus encodes UMP synthase, a bienzyme protein containing the last two enzyme activities of de novo pyrimidine biosynthesis: orotate phosphoribosyltransferase and orotidylate decarboxylase. This locus lies within chromosome region 93B. New mutations have been used to refine the 93B cytogenetic map and a chromosome walk has been executed to clone DNA from this region. DNA encoding UMP synthase was identified using mixed oligonucleotides which were based on sequences derived from conserved peptide tracts of the protein in other species. cDNA clones of the embryonic UMP synthase mRNA have been isolated and used to define the extent of genomic DNA sequences which encode the transcript. The embryonic RNA is approximately 1.75 kb in length.

Amino Acid Sequence↗
PubMed1990

Dangers of defibrillation: injuries to emergency personnel during patient resuscitation.

There have been no studies of the nature of defibrillatory shocks or risks to persons, providing them other than one individual case report and a number of incidental accounts. In this study, the severity and nature of injuries to prehospital emergency personnel in King County, Washington are reported. In addition, the types of injuries occurring to defibrillator operators throughout the United States, as voluntarily reported to the Food and Drug Administration (FDA), are described. In King County, prehospital emergency personnel reported eight accidental shocks. One individual was admitted to the hospital for 3 days and required lidocaine for premature ventricular contractions. Most injuries were the result of accidental contact with the patient rather than equipment failure, but the most serious case was caused by equipment failure. There were 13 injuries reported to the FDA over a 3 1/2-year period, and most injuries involved a mild shock or burn. Three patients were admitted to the hospital for observation. Two cases involved equipment failure. The rate of injury for paramedics was 1 per 1,700 defibrillatory shocks, and the rate of injury for emergency medical technician-defibrillator personnel was 1 per 1,000 defibrillatory shocks. These rates probably overestimate the real risk. Emphasis on safety and incorporation of safety procedures into resuscitation protocols can make the rate of injury even lower.

Allied Health Personnel↗
PubMed1990

Mobile element insertions causing mutations in the Drosophila suppressor of sable locus occur in DNase I hypersensitive subregions of 5'-transcribed nontranslated sequences.

The locations of 16 mobile element insertions causing mutations at the Drosophila suppressor of sable [su(s)] locus were determined by restriction mapping and DNA sequencing of the junction sites. The transposons causing the mutations are: P element (5 alleles), gypsy (3 alleles), 17.6, HMS Beagle, springer, Delta 88, prygun, Stalker, and a new mobile element which was named roamer (2 alleles). Four P element insertions occur in 5' nontranslated leader sequences, while the fifth P element and all 11 non-P elements inserted into the 2053 nucleotide, 5'-most intron that is spliced from the 5' nontranslated leader approximately 100 nucleotides upstream of the translation start. Fifteen of the 16 mobile elements inserted within a approximately 1900 nucleotide region that contains seven 100-200-nucleotide long DNase I-hypersensitive subregions that alternate with DNase I-resistant intervals of similar lengths. The locations of these 15 insertion sites correlate well with the roughly estimated locations of five of the DNase I-hypersensitive subregions. These findings suggest that the features of chromatin structure that accompany gene activation may also make the DNA susceptible to insertion of mobile elements.

Animals↗
PubMed1989

Predicted structures of apolipoprotein II mRNA constrained by nuclease and dimethyl sulfate reactivity: stable secondary structures occur predominantly in local domains via intraexonic base pairing.

Analyses of apolipoprotein II mRNA with chemical and enzymatic probes showed that double- and single-stranded regions were distributed uniformly along the mRNA except for a large (72 nucleotides) single-stranded region containing the translation stop codon. Secondary structure models constrained by the experimental data were made by varying the distance (along the mRNA) over which base pairing was allowed. Four prominent secondary structures were seen with restrictions of 165, 330, or 659 nucleotides suggesting that such structures from via local interactions over distances of 50-120 nucleotides. Predicted long range interactions involve only 2-3 base pairs while local interactions involve helices of 4-10 base pairs. Predicted helices of greater than or equal to 4 base pairs occur primarily within exons, raising the possibility that prominent secondary structures in mRNAs may be largely due to intraexonic base pairing. Tests of single- and double-stranded domains by oligonucleotide-directed RNase H cleavage and primer extension were in accord with the structure model and with nuclease and chemical modification data. The model predicting base pairing between the coding and the 3' noncoding regions was tested by RNase H cleavage followed by oligo(dT)-cellulose chromatography to separate 5' and 3' mRNA fragments. Most (82%) of the 5' fragment remained associated with the 3' noncoding region in a structure with a tm = 50 degrees C in 0.2 M Na+ suggesting that this stem could be stable in vivo. This stem may be stable in the isolated mRNA, but would likely occur transiently in polyribosomal apolipoprotein II mRNA due to ribosome transit through the 5' side of the stem. Alternate structures may occur in this region during ribosome transit and play a role in translation termination or in determining the susceptibility of the mRNA to degradation.

Animals↗
PubMed1989

Influence of abasic and anucleosidic sites on the stability, conformation, and melting behavior of a DNA duplex: correlations of thermodynamic and structural data.

We report a complete thermodynamic characterization of the impact of abasic and anucleosidic lesions on the stability, conformation, and melting behavior of a DNA duplex. The requisite thermodynamic data were obtained by using a combination of spectroscopic and calorimetric techniques to investigate helix-to-coil transitions in a family of DNA duplexes of the form d(CGCATGAGTACGC).d(GCGTACXCATGCG), where X corresponds to a thymidine residue in the parent Watson-Crick duplex and to an abasic or anucleosidic site in the modified duplexes. The data derived from these studies reveal that incorporation of an abasic site into a DNA duplex dramatically reduces the duplex stability, transition enthalpy, and transition entropy. The magnitudes of these lesion-induced effects are greater than one would expect based on simple nearest-neighbor considerations. Nearly identical thermodynamic data are obtained when the modified duplex contains an anucleosidic site rather than an abasic site. This observation suggests that the thermodynamic impact of these lesions primarily results from removal of the base rather than the sugar ring. Significantly, the melting cooperativities of the abasic and anucleosidic derivatives are identical with each other and with the corresponding unmodified Watson-Crick parent duplex. This result suggests that the phosphodiester backbone, rather than the base-sugar network, serves as the primary propagation path for the communication of cooperative melting effects. We propose molecular interpretations for the thermodynamic data based on the structural picture that has emerged from the NMR studies of Patel and coworkers on the same family of modified and unmodified DNA duplexes [Kalnik, M.W., Chang, C.-N., Grollman, A.P. & Patel, D.J. (1988) Biochemistry 27, 924-931].

Apurinic Acid↗
PubMed1989

Management of oesophageal stenosis in epidermolysis bullosa dystrophica.

Seven patients with epidermolysis bullosa dystrophica and chronic and recurrent oesophageal lesions such as spasm, strictures, and complete occlusion were studied. Dysphagia could be cured with drugs if it was caused by bullae formation or spasm. If oesophageal strictures were present, endoscopy and bouginage with corticosteroid prophylaxis during the quiescent phase of the disease was a safe and useful procedure. We have also given corticosteroids, which reduced the oedema caused by bullae formation and oral phenytoin, which reduced epithelial detachment by inhibiting collagenase activity. Verapamil counteracted oesophageal spasm and pureed food during periods of dysphagia reduced blistering of the upper oesophagus.

Adolescent↗
PubMed1988

Inactivation of chymotrypsin and human skin chymase: kinetics of time-dependent inhibition in the presence of substrate.

The reaction of chymase, a chymotryptic proteinase from human skin, and bovine pancreatic chymotrypsin with a number of time-dependent inhibitors has been studied. An integrated equation, relating product formation with time, has been derived for the reaction of enzymes with time-dependent inhibitors in the presence of substrate. This is based on a two-step model in which a rapidly reversible, non-covalent complex (EI) is formed prior to a tighter, less readily reversible complex (EI)*). The equation depends on the simplifying assumption [I] much greater than [E], but is applicable to reversible and irreversible slow-binding and tight-binding inhibitors whether or not they show saturation kinetics. The method has been applied to the reaction of chymase and chymotrypsin with the tetrapeptide aldehyde, chymostatin, basic pancreatic trypsin inhibitor and Ala-Ala-Phe-chloromethylketone (AAPCK). The irreversible inhibitor, AAPCK, showed the expected saturation kinetics for both enzymes and the apparent first-order rate constants (k2) and dissociation constants (Ki) for the non-covalent complexes were determined. Chymostatin was a much more potent inhibitor which failed to show a saturation effect. The second-order rate constant of inactivation (k2/Ki), the first-order reactivation rate constant (k-2), and the dissociation constant of the covalent complex (Ki*) were determined. Basic pancreatic trypsin inhibitor, a potent inhibitor of chymotrypsin, had similar kinetics to chymostatin but failed to inhibit chymase. The applicability of the two-step model and the integrated equation to slow- and tight-binding inhibitors is discussed in relation to a number of examples from the literature.

Amino Acid Chloromethyl Ketones↗
PubMed1988

Fluorescent probes of electrostatic potential 1 nm from the membrane surface.

We measured the electrostatic potential 1 nm from the surface of charged phospholipid bilayer membranes to test the predictions of the Gouy-Chapman theory. Fluorescent probes (anthraniloyl, 5-(dimethylamino)naphthalene-1-sulfonyl, Lucifer yellow) were attached covalently to the sialic acid residue of the ganglioside galactosyl-N-acetylgalactosaminyl(N-acetylneuraminyl)galactosylglucosylc eramide (GM1). These fluorescent gangliosides were incorporated into neutral [phosphatidylcholine (PC)] or charged [phosphatidylserine (PS)] phospholipid bilayers, and the fluorescence was quenched with the cations thallium and 4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl (tempamine). We calculated the electrostatic potential at the chromophore from the quenching ratio using the Boltzmann relation: the average potential was -30 mV for PS bilayers in 0.1 M NaNO3. We assume the chromophore is 1 nm from the surface because X-ray diffraction measurements demonstrate that the sialic acid residue of GM1 is 1 nm from the surface of a PC/GM1 bilayer [McDaniel, R. V., & McIntosh, T. J. (1986) Biophys. J. 49, 94-96]. We also used thallium and tempamine to quench the fluorescence of chromophores located at the surface of the PS membranes; in 0.1 M NaNO3 the average surface potential was -80 mV, which agrees with other measurements. The Gouy-Chapman theory predicts that the potential 1 nm from a membrane with a surface potential of -80 mV is -24 mV; this prediction agrees qualitatively with the experimental results obtained with fluorescent gangliosides.

Electrochemistry↗
PubMed1988

Residential mobility during pregnancy: implications for environmental teratogenesis.

Although most teratogens are suspected to act early in the first trimester of pregnancy, birth defects monitoring programs and etiologic studies usually use residence at birth as a proxy measure for residence in the first trimester in searching for environmental teratogens. Because of the high mobility of the U.S. population, residence misclassification can potentially alter inferences concerning environmental teratogens. To evaluate this potential bias, data from the population-based Maryland Birth Defects Reporting and Information System were analyzed. In 1984, the system ascertained 295 infants with one or more of 12 sentinel defects. Of these cases, 59 (20%) mothers reported they have changed address between the time of conception and the time of birth, and 22 have moved to a different county. The residential mobility rate varied by demographic variables and was highest among white women, in the age group 20-24 years. If residence at birth is used as a screening test for residence at conception, it can be shown that in the presence of an environmental teratogenic exposure, misclassification of exposure increases with increasing mobility rate, and population exposure frequency. Such misclassification tends to weaken associations between residence and birth defects and may lead to missing environmental teratogens. This analysis emphasizes the need to use residence information early in pregnancy rather than exclusively at birth.

Abnormalities, Drug-Induced↗