Gunther L. Eichhorn

DNA methylation in aging of mice

Methylation of cytidine residues of DNA appears to be involved in the control of gene expression; therefore, hypomethylation of the DNA can be considered to be an active rather than a passive process. Previous studies of mammalian DNA methylation during aging have produced an assortment of results. In this study, we have examined the change in DNA base composition, including the change in 5-methyldeoxycytidine (m5dC) contents with age of mice. Livers pooled from 6 mice from each of six age groups between 6 and 31 months have been subjected to a sensitive analytical technique (HPLC). The DNA composition of different age groups is very consistent in most aspects. The ratio of (dA + dT)/(dG + dC + m5dC) as well as the sum of dC and m5dC remain constant throughout the animals lifespan. However, a consistent gradual decline in m5dC content is noted as the age increases to 24 months. Thus, the 6-month-old animal pool exhibits the largest amount of m5dC (1.67 +/- 0.2%), which is reduced consistently as the animals age reaches 24 months. This decrease in m5dC is accompanied by an increase in dC. No further decrease in m5dC occurs after 24 months; in fact, the data could indicate an increase after that age. No dTs are apparently produced by deamination of m5dC.

The interaction of metal ions with polynucleotides and related compounds: VI. Effect of metal ions on the anion-exchange behavior of the nucleotides☆

The activity of ribonuclease increases with increasing concentration of added divalent metal ions until the attainment of an optimal metal concentration, beyond which the addition of further metal ions inhibits the reaction. When either the optimal metal concentration or the enzymatic activity at that concentration is plotted versus the atomic number of the transition metals, the resulting curves can be correlated with the stability constants of complexes of the metals. These results are in line with metal ions reacting at two types of sites, one activating and the other inhibiting. Crystalline bovine pancreatic ribonuclease is affected in the same way as ribonuclease A. The activation of deoxyribonuclease I has been reinvestigated. Metal ions generally activate this enzyme, but the relative activating ability of the various metal ions is not readily correlated with complexing ability, as in the case of ribonuclease. Co(II) activates deoxyribonuclease I better than the metals that have generally been used for this purpose.

Interaction of metal ions with polynucleotides and related compounds. XX. Control of the conformation of polyriboadenylic acid by divalent metal ions

Abstract Divalent metal ions have profound effects on the conformation of poly A, as indicated by changes in optical rotatory dispersion and ultraviolet spectra, with different metal ions having distinctly different effects. At pH 6 and in the presence of 5 × 10 −2 M NaCl, two moles Ni(II) or Co(II) per mole of poly A residue stabilize the single helix; the same amount of Mg(II), Ca(II), Zn(II) or Mn(II) produce a mixture of double and single helix, while Cu(II) and Cd(II) induce the formation of a random coil structure. Under other conditions, these same metals have different effects, with the amount of added NaCl an important factor that can be used to control the way in which a particular divalent metal ion will induce the transition from a double helix to single helix or from helix to coil. The single helix is stabilized by metals binding to phosphate; the double helix is destabilized by metal ions binding to either phosphate or base, the former producing single helix and the latter random coil. The melting temperature of double helical poly A is diminished when metal ions bind to either type of site.

Physical chemical studies on the age changes in rat tail tendon collagen

Abstract Collagen was solubilized from the tail tendons of rats of different ages in dilute acetic acid. Optical rotation measurements on the solubilized collagen indicated that no significant age changes occur in the overall secondary structure of collagen. Sedimentation velocity measurements on the solubilized collagen in KSCN solutions, in which the secondary structure is broken down, revealed only α (single-stranded) and β (double-stranded) subunits. The portion of α in the acid-solubilized collagen, measured by sedimentation velocity, decreases markedly between 1 and 3 months but remains fairly constant thereafter. The rate of solubilization and the fraction of the total collagen eventually solubilized, however, showed drastic changes after 3 months, but not before. These observations are in line with a continuing production of covalent crosslinks in the collagen fiber throughout the life span of the animal, according to the following kind of picture: β subunits are, effectively, first produced by crosslinking α subunits within the tropocollagen molecules laid down in the formation of the fiber. Crosslinking then spreads randomly within and between tropocollagen units, and eventually an insoluble matrix is built up, so that less and less solubilizable collagen becomes more and more entrapped.

Polynucleotide cross-linking by aluminum

The observations that there was an increased concentration of Al in the brains of Alzheimers, Guam-Parkinson, and amyotrophic lateral sclerosis disease patients and that there was an apparent localization of the Al in chromatin led to a study of the interaction of Al(III) with DNA. We have previously shown that Al cross-links calf thymus DNA at low pH (S. J. Karlik, G. L. Eichhorn, P. N. Lewis, and D. R. Crapper, Biochemistry 19, 5991 [1980]). Extended studies indicate that cross-linking occurs in DNAs of all base ratios, including polydAdT and polydGdC. Since Al cross-links prevent renaturation in polydAdT, the decrease in the amount of polymer renatured in the presence of Al becomes a quantitative appraisal of the extent of cross-linking. Saturation of cross-linking occurs at a 0.4 ratio of Al to nucleotide phosphate, indicating that potentially 80% of the base pairs are Al bound. Cross-links are broken at elevated pH and by EDTA.

Is there any relationship between aluminum and Alzheimer's disease?

The controversial role of aluminum in Alzheimers disease (AD) is reviewed. While current data would suggest the lack of a causative role, alterations in the brain and other organ systems caused by AD might increase the penetration of aluminum as well as other metals into the brain and lead to their contribution to such pathological features as neurofibrillar tangles (NFTs).

Conformational change induced by metal ions through coordination

Abstract The structure of a metal complex may be determined by the structure of the ligand or by the electronic configuration of the metal. In the latter case, the metal can induce a conformational change in the ligand. Such a conformational change is a usual result in coordination reactions, and is particularly significant in biological macromolecules, such as nucleic acids, proteins, and enzymes. We have recently obtained evidence for a mechanism that involves a metal participating in conformational changes that occur in both enzyme and substrate simultaneously.

Some effects of metal ions on DNA structure and genetic information transfer

The reaction of metal ions with nucleic acids can lead to a variety of dramatic effects on nucleic acid structure, e.g., crosslinking of the polymer strands, degradation to oligomers and monomers, stabilization or destabilization, and the mispairing of bases. These effects have important implications for genetic information transfer. Metal ions are involved in many aspects of this transfer; we are presently concerned with the effect of metal ions on the orientation of the active site of RNA polymerase.Many of the effects of metal ions on nucleic acid structure involve changes in the conformation of the macromolecules. We have found that conditions that have been used to convert B DNA to Z DNA lead to at least two other conformational changes, and phase diagrams delineate the realms of stability of each of the forms. We have carried out a number of studies that demonstrate that the conversion of B to Z DNA is very closely correlated with a substantial decrease in the ability of the DNA to act as a template for RNA synthesis.

The effect of divalent ions on the isolation of proteins from rat liver nucleoprotein

Abstract The nature and quantity of acid-soluble proteins separated from rat liver nucleoprotein is greatly dependent upon the presence of divalent metal ions. In the absence of these ions, and particularly in the presence of EDTA, low yields of protein are obtained because of greater solubility of the nuclear contents. Moreover, the use of EDTA selectively removes the “arginine-rich” proteins from the nucleoprotein pellet, so that eventual protein fractionation results in relatively extensive retention of “arginine-poor” proteins. The use of divalent metal ions (Mg 2+ , Mn 2+ ) produces similar fractionation patterns. The use of NaCl as the electrolyte allows the isolation of some arginine-rich protein, but in very low yield when compared to the preparations using divalent ions. Thus, ionic strength is a minor factor in the extraction of the arginine-rich proteins from such nuclei, but the presence of divalent metal ions is the most important requirement. Evidently these metal ions exercise an important role in maintaining the structural integrity of arginine-rich nucleoproteins in the nuclei.

A simple probe for DNA accessibility in chromatin.

When DNA is treated with Cu(II) and then heated, the melting temperature (Tm) of the DNA is dramatically decreased (8). The Cu(II) binds to the DNA in such a way as to destabilize the double helix and help to break the hydrogen bonds between the bases. When soluble chromatin is similarly treated with Cu(II) and heated, the Tm is unaffected. Apparently the Cu(II) cannot penetrate the chromatin structure and thus cannot initiate the DNA destabilization process. However, when H-1 histone is removed from the chromatin by affinity chromatography, subsequent treatment with Cu(II) does lead to a lowered Tm when the chromatin is heated. This Tm lowering is also achieved by two less drastic techniques that do not remove histone H-1, but decrease the affinity of the H-1 to the DNA: (1) a mild acetylation procedure that specifically modifies either 2 or 4 epsilon-amino groups of lysines on the H-1 histone, and (2) reaction with phosphate-binding divalent metal ions, e.g., Mg(II), Mn(II), or Co(II). Apparently, removal of H-1 or decreased affinity of H-1 for DNA increases the accessibility of the DNA to the Cu(II). This phenomenon suggests a very simple qualitative probe for the degree of structural change in chromatin produced by a change in the stability of the DNA-H-1 interaction.