Wednesday, June 24, 2009

Response to Zhang: another trick of heavy isotopes

Deuterium cannot be kept for long enough at
exchangeable DNA sites to achieve this desired effect in
vivo because it will readily (within a few minutes) be
replaced by hydrogen atoms from normal water, the major
component of live cells, owing to fluctuational openings
of DNA base pairs (DNA ‘breathing’). Otherwise,
normal water would have to be almost completely replaced
by heavy water, which is not possible owing to the
known lethal toxicity of heavy water for mammals at
concentrations >35%. This was the reason that high
KIE was only observed in the condensed phase or in waterfree
solutions whereas even traces of natural water basically
abolished this effect.
For this reason, to achieve the goal of reducing the
amount of unfavorable tautomers it was suggested that
deuterium be placed at non-exchangeable sites of DNA
purines and pyrimidines. In this case, other heavy
isotopes can also be used for KIE (Figure).

caution should be exercised in choosing sites for isotopic
replacement because certain studies estimated an unusually
highamount of unfavorable tautomers in tRNAwhen
the natural 14N isotope was completely replaced with its
heavier 15Nsubstitute.Note that thishighamount of
unfavorable tautomers exceeds the amount of mutationcausing
tautomer of a highly mutagenic analog of cytosine. It is noteworthy that isotopic changes in non-exchangeable
DNA sites might reduce the level of age-related
mutations, including epigenomic modifications, by other
mechanisms too.

Vadim V. Demidov
Trends in Biotechnology
Volume 26, Issue 3, March 2008

Wednesday, June 17, 2009

Another trick of heavy isotopes

Hong-Yu Zhang speculate that non-radioactive heavy
isotopes might resist diseases and ageing by another
Forty years ago, Lo¨wdin proposed a DNA mutation
model that was built on inter-base double proton (or hydrogen)
transfer (DPT).

As illustrated in Figure, following the DPT the four bases (A, G, C, T) are turned into
tautomeric forms (A*, G*, C*, T*), which will no longer
bind to the normal Watson–Crick partners but to others,
especially C, T, A and G, respectively, resulting in new base
pairs: A*–C, G*–T, C*–A and T*–G. As a result, nucleotide
mutation occurs, which results in a time-dependent loss of
genetic information and thus leads to various diseases and
ageing. Since the first description of the model it has
attracted much interest and has been supported by
increasing theory and experimental evidence.
According to the principle of KIE, the inter-base DPT
can be efficiently slowed down by substituting the transferred
protons with heavy isotopes. For instance, a high
KIE (7.4) for deuteration has been observed in photoinduced
inter-base DPT of model DNA base pairs.
Therefore, we propose that by replacing the transferred
hydrogen atoms (labeled in red in Figure) in the base
pairs with deuterium atoms, the DPT rate will be considerably
reduced, which should reduce the possibility
for DNA mutation and thus slow the progression of
some diseases, as well as the ageing process. Consequently,
we suggest that another way for heavy
isotope substitution in biomolecules to benefit our health
exists – that is, through decreasing the DPT-based DNA
mutation rate.

Another trick of heavy isotopes
Hong-Yu Zhang
Trends in Biotechnology
Volume 26, Issue 3, March 2008, Pages 118-120

Wednesday, June 10, 2009

Deuteronation and aging.

Deuterium has one proton and one neutron in its atomic nucleus, but hydrogen has only proton. The natural abundance of deuterium is 1 per approximately 6600 hydrogen atoms. Therefore deuterated water (both HOD + D(2)O [heavy water]) abundance is 1 per approximately 3300 water molecules. One dissociation product of deuterated and heavy water is deuteron (proton + neutron, D(+), H(2)OD(+)/D(3)O(+)). Because heavy water has a lower ionization constant than water, the D(+)/H(+) ratio is approximately 1/15,000 in biological fluids. O-D bond length is shorter than O-H, and D-O-D angle is lesser than H-O-H. Once a deuteron exchanges with proton on the water-exposed surface of a macromolecule, it can lead to a conformational change and the reverse exchange will be less likely. Deuteron bonds are stronger than proton bonds. Therefore an increase of deuteronated macromolecules can be expected in due course of time. In order to test this hypothesis, we conducted a pilot study and measured the D/H ratio in the tails of three Sprague-Dawley rats at different ages (4 weeks, 5 weeks, and >1-year old) by elemental analysis coupled with isotope ratio mass spectrometry (EA-IRMS) technique. To prevent the effect of daily water consumption, the homogenized tails were lyophilized before analysis. The results, as mean of several measurements, of 4 weeks, 5 weeks, and >1-year-old rats were per thousand-94 +/- 9.56, per thousand-101.71 +/- 6.89, per thousand-83.68 +/- 3.46 delta((2)H) relative to VSMOW, respectively. Although there is a slight increase in >1-year-old rat, the difference among the animals was not significant. We propose that, before reaching to a final conclusion about the accumulation of deuterium with aging, the measurements should be done not in whole tissue samples but in purified macromolecules from a larger set of animals.

Ann N Y Acad Sci. 2007 Apr;1100:400-3.
Olgun A, Oztürk K, Bayir S, Akman S, Erbil MK.

Wednesday, June 3, 2009

New anti-ageing strategy on course: Kinetik Isotope Effect vs. ROS

Oxford-based researcher Mikhail Shchepinov noticed that ROS oxidize cellular substrates by a mechanism typically involving hydrogen abstraction in a rate-limiting step. Through his university background in chemical kinetics, Shchepinov also knew about the effect of an isotope substitution on a rate of chemical transformation. The strength of a covalent bond is subtly influenced by the atomic masses at either end of the bond: heavier isotopes form stronger bonds than light isotopes of the same elements, and so reactions involving the breaking of those bonds proceed more slowly. For example, hydrogen comes in two stable isotopes, the common ‘light’ hydrogen isotope (H) and its twice-heavier sibling, deuterium (D). The C–D bond is significantly stronger than the C–H bond, and therefore a cleavage of the former bond will occur several times more slowly than the corresponding cleavage of the latter bond. The common ‘light’ carbon atom 12C of the C–H bond can also be substituted for a heavier, stable 13C isotope, but the bond-cleavage-rate decrease will be smaller than that involving substitution of H for D because 13C is only 8% heavier than 12C. This is called the kinetic isotope effect (KIE).

Shchepinov's idea is to use the KIE to make proteins, nucleic acids and lipids more resistant to ROS damage. It is summarized as follows: replace all ‘light’ atoms at the ROS-targeted sites of biomolecules with their heavier isotopes, which should evidently be both non-radioactive (stable) so as not to damage the cell and non-exchangeable with the intracellular environment to avoid their loss via exchange with normal ‘light’ isotopes. Besides non-radioactive D and 13C, stable 15N and 18O ‘heavy’ isotopes can also be used to take the place of common ‘light’ nitrogen and oxygen constituents of biomolecules. Note that such isotopic ‘make-up’ will not change the chemical nature of the compounds: the bonding structure of isotopical ‘twins’ will be identical, and their chemistry will basically be similar. However, the heavier sibling will be less reactive. Also note that even if the ‘heavy’ isotope is not a part of the bond that is cleaved by ROS, a small decrease in the oxidation rate, known as a secondary KIE, can nevertheless usually be observed.

Vadim V. Demidov (2007) Heavy isotopes to avert ageing?
Trends in Biotechnology, Volume 25, Issue 9, September 2007, Pages 371-375

Current defense strategies from reactive oxygen species (ROS)

The traditional strategy of combating free radicals is based on counteracting ROS with antioxidants, free-radical scavengers or readily oxidizable substances acting as free-radical captors. Common antioxidants include vitamins (vitamin C, vitamin E), carotenoids (β-carotene, lycopene) and microelements (zinc and selenium; the latter can also enhance the body's own repair pathways). To prevent ageing and age-related diseases, these compounds are usually taken as dietary supplements, which are widely used in industrialized countries.

It is thought that such nutraceuticals protect human cells and tissues from the damaging effects of oxidation. Many nutraceutical and health-food companies are now involved in the production and distribution of antioxidant-containing dietary supplements. However, despite widespread public optimism about the likely beneficial anti-ageing effects of antioxidants, so far there is no solid proof that use of antioxidants can stop or reverse the ageing process. Although some researchers claim that antioxidant supplements do show positive results in the battle against ageing, others have not identified any statistically significant benefit from the use of antioxidants. Some studies indicate that dietary antioxidants might only be effective in men because their bodies are somewhat lower in the contents of certain antioxidants, for example, β-carotene, than women's bodies.

However, the amounts in which these antioxidants should be consumed to be truly beneficial without being harmful has to be carefully considered, especially for elderly or other vulnerable populations. Indeed, by modulating cellular redox status, an optimal ROS level could be very important in homeostasis of aerobic life. In this regard, a recent study has come to a shocking conclusion: treatment with β-carotene, vitamin A and vitamin E might actually increase mortality! This study also concluded that the potential roles of vitamin C and selenium in longevity in humans need further investigation.

The contradictory results obtained with antioxidants as anti-ageing agents make it clear that alternative approaches are needed. A fresh look at the old problem has just been taken from the chemical kinetics viewpoint.

Vadim V. Demidov (2007) Heavy isotopes to avert ageing?
Trends in Biotechnology, Volume 25, Issue 9, September 2007, Pages 371-375