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Biomedical subjects

M J Behe

Publications and source records attributed to M J Behe.

35 records · Page 2Linked to original sources
PubMed1988

Sequence dependence of DNA structure. The B, Z, and A conformations of polydeoxynucleotides containing repeating units of 6 to 16 base pairs.

In order to study the sequence dependence of the B-Z transition, we have synthesized eight double-stranded polydeoxynucleotides. The polymers have defined alternating purine-pyrimidine sequences with repeating units of 6-16 base pairs, and contain 12.5-33% AT base pairs. A and T nucleotides are in nearest-neighbor positions in the series poly[d[TA(CG)2-7]], but are isolated from each other, with minimum separations of 2 and 6 GC base pairs, in poly[d[TGCA(CG)6]] and poly[T(GC)3A(CG)4)]], respectively. All of the polymers except poly[d[TA(CG)2]] were shown by circular dichroism to undergo a right- to left-hand helical transition at high NaCl concentrations, and all polymers exhibited a B to A transition in the presence of ethanol. Poly[d[TA(CG)7]] was the only polymer to undergo a B to Z to A transition in ethanol. At a constant percentage of AT base pairs, the B-Z transition is sequence-dependent, occurring at lower salt concentrations for polymers containing longer runs of contiguous GC base pairs in the repeating unit.

Chemical Phenomena↗
PubMed1988

An oligopurine sequence bias occurs in eukaryotic viruses.

Twenty four DNA and RNA viral nucleotide sequences, comprising over 346 kilobases, have been analyzed for the occurrence of strings of contiguous purine or pyrimidine residues. On average strings greater than or equal to 10 contiguous purines or pyrimidines are found three and a half times more frequently than would be expected for a random distribution of bases. Detailed analysis of the 172 kilobase Epstein-Barr viral sequence shows that the bias in favor of contiguous purine residues increases with the length of the purine string. These findings are similar to those seen for genomic DNA from higher eukaryotes. In contrast no overrepresentation of oligopurine or oligopyrimidine strings is observed in 52 kilobases from eight bacteriophage and E. coli DNA sequences.

Base Sequence↗
PubMed1988

Purification and properties of an acid phosphoprotein phosphatase from lactating bovine mammary gland with activity toward phosphotyrosine.

An acid phosphatase has been partially purified from lactating bovine mammary gland. Properties of this enzyme were compared with those of a well-characterized phosphoprotein phosphatase from bovine spleen. The two enzymes were similar in their activation by sulfhydryl reagents and inhibition by metal chelating agents. Both enzymes rapidly hydrolyze ATP and aromatic phosphates and are relatively inactive toward alkyl phosphates; both are tartrate-resistant phosphatases. The mammary enzyme has a low Michaelis constant for alpha s1-casein (42 microM), and thus, like the spleen enzyme, appears to be a phosphoprotein phosphatase. Finally, the spleen and mammary enzymes displayed reactivity toward phosphotyrosine, a model substrate for phosphotyrosyl protein phosphatase. Thus, the phosphatases from spleen and mammary gland are quite similar in reactivity and could possibly be similar in function.

Acid Phosphatase↗
PubMed1987

The DNA sequence of the human beta-globin region is strongly biased in favor of long strings of contiguous purine or pyrimidine residues.

The DNA sequence of the human beta-globin region, comprising over 67 kilobase pairs, has been analyzed for the occurrence of strings of contiguous purine or pyrimidine residues. Tracts of 10 or more contiguous residues are found 4 times more frequently than would be expected with a random distribution of bases, so that a long string occurs at an average of every 250 base pairs. A survey of six other human gene sequences, totaling 86 kilobase pairs, shows a remarkably similar result. No such overrepresentation of contiguous purine or pyrimidine residues is found in the bacteriophages lambda or T7.

Base Sequence↗
PubMed1987

Influence of tetraalkyl ammonium ions on the structure of poly (rG-dC).poly (rG-dC): unexpected transitions among the Z, A and B conformations.

The conformation of the double-stranded, mixed ribodeoxyribo polynucleotide, poly (rG-dC).poly (rG-dC), has been examined in the presence of tetraalkyl ammonium ions. Tetramethyl ammonium ion stabilizes the "low salt" Z conformation (1) of the polymer from submillimolar to molar concentrations of the counterion. In the presence of tetraethyl and tetrapropyl ammonium ions the polymer exists in the low salt Z form up to 2 mM concentration of the counterions and then flips to the right hand helical A form. With tetrabutyl ammonium counterions the polymer is in an A conformation at low ion concentrations and converts to a B form at concentrations greater than thirty millimolar. These results are interpreted in terms of electrostatic and solvent interactions of the polynucleotide.

Chemical Phenomena↗
PubMed1987

Binding of p-nitrophenyl phosphate and other aromatic compounds by beta-lactoglobulin.

Results obtained from gel filtration showed that beta-lactoglobulin binds p-nitrophenyl phosphate with a stoichiometry of 1 mol of ligand per 18,360 monomer. Circular dichroic spectra confirmed the binding and implicated tryptophan and phenylalanine residues in the interaction. Fluorescence of the protein was quenched on binding also supporting complex formation; analysis of these data indicates that p-nitrophenyl phosphate binds to beta-lactoglobulin A with a dissociation constant of 31 microM. The B and C genetic variants of beta-lactoglobulin bind p-nitrophenyl phosphate with dissociation constants of 63 and 70 microM, respectively. In addition, a series of other nitrophenyl compounds and pyridoxal phosphate were also investigated by fluorescence analysis and found to bind to the protein. These results are discussed with respect to a recent hypothesis that beta-lactoglobulin binds retinol and is structurally related to serum retinol binding protein.

Lactoglobulins↗
PubMed1986

The B-Z transition of a polynucleotide with a 10-base pair repeating sequence.

In this communication we report the synthesis and characterization of a defined-sequence, alternating purine-pyrimidine polymer with a 10-base pair repeating unit: poly[d(CGCGCGTGCA)]. It is seen that the polymer can undergo a right-left-hand helical transition, but the conformational transition requires much higher salt concentration than with poly[d(G-C)]. The results are interpreted in terms of interaction of the polymer with solvent molecules.

Circular Dichroism↗
PubMed1985

Methylated pyrimidines stabilize an alternating conformation of poly(dA-dU).poly(dA-dU).

We have investigated the effect of increasing percentages of methylated pyrimidines on the structure of poly(dA-dU).poly(dA-dU). This was done by synthesizing analogous polynucleotides that contained deoxythymidine residues as well as deoxyuridine residues and observing their 31P NMR spectra in increasing amounts of CsF. The results show that methylated pyrimidines play a large role in the stabilization of the "alternating B" conformation of DNA.

Magnetic Resonance Spectroscopy↗
PubMed1985

Salt induced transitions between multiple conformations of poly (rG-m5dC).poly (rG-m5dC).

Salt induced transitions between four conformations of the methylated ribo-deoxyribo co-polymer poly (rG-m5dC).poly (rG-m5dC) have been studied using phosphorous-NMR, Raman spectroscopy, and circular dichroism. A high salt A-Z transition is observed for the polymer. However, the methylated polymer does not enter the high salt Z form more readily than the analogous unmethylated polymer, unlike the effect of methylation on the fully deoxy polymer poly (dG-dC).poly (dG-dC). The methylated polymer fails to undergo a low salt A-Z transition in 5 mM Tris buffer, unlike the unmethylated poly (rG-dC).poly (rG-dC). However, if the counterion is changed to triethanolamine buffer, an A-Z transition does take place. In 5 mM Tris buffer the phosphorous-NMR spectrum of poly (rG-m5dC).poly (rG-m5dC) shows one resonance in the absence of NaCl that splits into two closely spaced resonances as the NaCl level is increased to 30 mM. The Raman spectrum of poly (rG-m5dC).poly (rG-m5dC) shows that it is in the A conformation at intermediate salt concentrations. From this we conclude that poly (rG-m5dC).poly (rG-m5dC) is in a regular A conformation in Tris buffer at low Na+ levels, shifting to an alternating A conformation with a dinucleotide repeat at intermediate salt concentrations.

Circular Dichroism↗
PubMed1984

Critical amount of oligovalent ion binding required for the B-Z transition of poly (dG-m5dC).

By working at very low Na+ concentrations (1mM and less), the number of bound Mg(2+), cobalt hexamine (3+), and spermine (4+) necessary to induce the B-Z transition of poly (dG-m5dC) has been directly measured. The results show that if as little as 1 cobalt hexamine(3+) or spermine(4+) is bound per 40-50 nucleotides the transition will occur. A greater fraction of bound Mg(2+) is required, 1 bound Mg(2+)/10 nucleotides. The dependence of the transition midpoint concentrations of oligovalent ions on Na+ concentrations suggests that specific ion binding energies, not included in counterion condensation theory, are responsible for the transition.

Cations↗
PubMed1984

Salt-induced Z-A-Z transition sequence in the mixed ribo-deoxyribo copolymer poly(rG-dC) X poly(rG-dC).

The left-handed helical conformation of various polynucleotides has been seen in solution at higher salt concentration than has the right-handed helical conformation of the corresponding polymer. We report here, however, studies of conformational transitions in the mixed polymer poly-(rG-dC) X poly(rG-dC). This polynucleotide appears to exist in a Z conformation under low-salt conditions, converts to an A helix at intermediate salt, and reverts to a left-handed helix at high-salt concentration.

Circular Dichroism↗
PubMed1979

Quantitative assessment of the noncovalent inhibition of sickle hemoglobin gelation by phenyl derivatives and other known agents.

The ability of a variety of phenyl derivatives to inhibit sickle cell hemoglobin gelation was placed on a quantitative scale by parallel equilibrium and kinetic assays. Modifications of the phenyl ring studied include polar, nonpolar, and charged substituents, added aromatic rings, and loss of aromaticity. Other noncovalent inhibitors previously reported to have high potency were measured and placed on the same quantitative scale. Some phenyl derivatives were found to be as effective an any other known noncovalent antigelling agent. The phenyl compounds penetrate easily into red cells, and their potency is tolerant to chemical modification, which holds out the possibility of designing low-toxicity derivatives. On the negative side, the level of potency obtainable appears to be inadequate for clinical use. The best phenyl inhibitors display a functionally defined inhibitory constant (K1) of 75 mM, and it can be estimated that inhibitor concentrations over 20 mM would be necessary to obtain minimal clinically significant benefit. Furthermore, with the variety of modifications tested here, no impressive increase in activity could be achieved over that found in the simplest phenyl compounds.

Benzene Derivatives↗
PubMed1978

Sickle hemoglobin gelation. Reaction order and critical nucleus size.

Sickle hemoglobin (Hb S) gelation displays kinetics consistent with a rate-limiting nucleation step. The approximate size of the critical nucleus can be inferred from the order of the reaction with respect to Hb S activity, but determination of the reaction order is complicated by the fact that Hb S activity is substantially different from Hb S concentration at the high protein concentrations required for gelation. Equilibrium and kinetic experiments on Hb S gelation were designed to evaluate the relative activity coefficient of Hb S as a function of concentration. These experiments used non-Hb S proteins to mimic, and thus evaluate, the effect on activity coefficients of increasing Hb S concentration. At Hb S concentrations near 20% the change in Hb S activity coefficient generates two-thirds of the apparent dependence of nucleation rate on Hb S concentration. When this effect is explicitly accounted for, the nucleation reaction is seen to be approximately 10th-order with respect to effective number concentration of Hb S. The closeness of the reaction order to the number of strands in models of Hb S fibers suggests a nucleus close to the size of one turn of the Hb S fiber. These experiments introduce a new approach to the study of Hb S gelation, the equal activity isotherm, used here also to show that Hb S.Hb A (normal adult hemoglobin) hybrids do incorporate into growing nuclei and stable microtubules but that A.S hybridization is neutral with respect to promotion of Hb S nucleation and the sol-gel equilibrium.

Chemical Phenomena↗