Can DNA repair enzymes reverse oxidative damage and combat aging?

[bioquest]

Are there any known enzymes that will fix oxidative dna damage without correcting mismatched bases? Any enzymes that are thought to work like that? thanks

 

[Ygggdrasil]

Yes. Photolyase is one such example that fixes thymine dimers (by using the energy from light which creates thymine dimers in the first place. Pretty cool!). Another example is 8-oxoguanine glycosylase which repairs oxidized guanine nucleotides. I'm sure there are other examples, but these are the first two on the top of my head.

 

[bioquest]

well I read thatPhotolyase is present and functional in prokaryotes, is present in lower eukaryotes (as yeast) where it is thought to have a minor role, and it has not been found in human cells. However, many higher eukaryotes, including humans, possesses a homologous protein called cryptochrome that is involved in light-sensitive regulatory activities such as modulating circadian rhythms.

but even though it's not found in human cells it could be introduced into them, through genetic manipulation or endosomes or something right, I mean could it be?

 

[Ygggdrasil]

Probably. You'd probably have to modify the enzyme to tell human cells to send the protein into the nucleus (remember prokaryotes have no nucleus so they lack nuclear localization signals that target them to the nucleus), but in theory there should be nothing that would prevent you from getting these proteins to work in humans.

 

[bioquest]

I know photolyse won't work; What are the other enzymes that would fix oxidative damage but not mismatched bases? 8-oxoguanine glycosylase can't fix all the oxidative dna damage right?

 

[Mike H]

8-oxoguanine glycosylase is a member of the larger DNA glycosylase family, which contains a number of related enzymes that do similar chemistry on different base pairs. There's even a uracil-DNA glycosylase, which sounded a bit strange to my ear the first time I heard of it, but it does exist.

Nucleotide excision repair (a more general and flexible alternative to more specific DNA repair mechanisms, as I recall) involves a number of proteins in eukaryotes, of which I can remember about two, if I'm lucky, on a good day. You may be interested in investigating that more carefully, as I am of no use on the subject.

 

[bioquest]

what about the enzymes in this plant 4767-year-old bristlecone pine although I heard some of the cells in it were only a couple of years old

 

[bioquest]

okay I just realized only the bark was alive in that plant or something/that I was misinformed but

Anyone have insight on what DNA repair enzymes/systems you would add to something to extend it's longevity/help with aging. overexpressing some of them seemed to cause more problems with mismatched bases, so I wanted to know about potential non human dna repair enzymes/systems that could be added to a mouse

 

[Mike H]

Many of these mechanisms are found in a number of organisms across all domains of life. They are not exactly the same, of course - for instance, nucleotide excision repair (NER) in eukaryotes involves more proteins than in prokaryotes - but the basic notion of NER is conserved.

I'm not aware of anyone trying to add a completely new DNA repair pathway to an organism, but DNA repair research is a rather large world to keep track of, especially if you're not in the field (such as myself). You might find the http://asajj.roswellpark.org/huberman/DNA_Repair/DNA_Repair.htm" site at the Roswell Park Cancer Institute useful as a starting point for navigating through the field. I would suggest starting with the references there, at least with regard to finding people who are currently involved with the cutting edge research and go from there.

Good luck!

 

[bioquest]

Does photolyse fix oxidative dna damage?

 

[Mike H]

The links provided have very clear explanations of what photolyase does, far more so than what someone who is not a specialist (aka me) can provide. The page on direct repair of DNA damage on the Roswell Park website mentions something that is of relevance to your earlier question about introducing a foreign photolyase to an organism. It takes a simple Google search to find the (freely available) paper which discusses the answer to this question.

I get the impression that we're being asked to spoon-feed homework answers. It may be uncouth to say so, but I can't shake the feeling...

 

[bioquest]

well but what I mean is, could photolyse solve the oxidative dna damage problem, or would it only fix some of the damage? I'm not in school yet so its not homework answers I mean I know it's called the homework help forums

 

[Ygggdrasil]

There are many types of oxidative damage that can occur on DNA. As such, there are many enzymes to recognize and fix oxidative damage. Photolyase will fix only a certain type of oxidative DNA damage (thymine dimers) and will not fix other types.

 

[bioquest]

Is there any known one enzyme that would repair all oxidative dna damage/is there any known combination of enzymes that would solve the oxidative dna damage problem if overexpressed or something? (Without causing higher problems in mismatched bases) What are the ones, if any, that would do it without fixing or attempting to fix/making mistakes in fixing mismatched bases?

ty

 

[Mike H]

Base excision repair mechanisms typically rely upon specific DNA glycosylases for proper functioning, so they're hardly an extremely general mechanism. The direct repair methods are usually only there for particular types of DNA damage (such as photolyase), so they will only revert back a particular sort of damage. Nucleotide excision repair relies upon structural perturbations to the DNA upon damage, so it's a bit more general, but it is not foolproof either. There are other mechanisms which come into play under certain circumstances, but are not otherwise active.

There's no one single DNA repair mechanism that I'm aware of that can repair everything with the necessary fidelity to the original genome. This is why there are so many different systems -even mismatch repair - to overlap one another and make sure that your genes stay intact.

 

[bioquest]

Is there any known thing- any combination or anything of enzymes etc I say combination because you said "there's no one single DNA repair mechanism that can repair everything with the necessary fidelity to the original genome" that if overexpressed or something or just used the way it is would solve/almost solve or anything the oxidative dna problem? without creating a higher problem with mismatched bases? (Since apparently it would see problems that weren't there and try to fix what didn't need to be fixed) I mean I guess it could/would cause other problems I was just asking about the ones that wouldn't cause increased problems with mismatched bases

I mean, are the answers to these no/unknown? with our current knowledge?

So direct reversal methods wouldn't solve the oxidative dna damage problem or anything, they would just work on some types of oxidative damage?

So there's not a group of enzymes, with for example something like photolyse in it, that could solve/mostly solve or anything the oxidative dna damage issue and not cause more problems with mismatched bases?

 

[Mike H]

From the little I've read since undergrad molecular biology, one would probably have to find a way to upregulate (overexpress) EVERY repair mechanism in order to ensure that adequate fidelity is ensured. The question is, given the baseline rate of mutations in eukaryotes (which is a value I don't remember), just how much upregulation would be required to suppress even that? If you need a huge jump in upregulation for a minimal effect in taking care of baseline mutation, it might not be worth it, which will make sense by the end of this post. I get the impression it could be one of those things where 10% effort will get you 90% of the desired results, but that last 10% of the desired results is extremely costly.

Another issue is how to actually initiate DNA repair mechanisms. I know that people have suggested that there are redox chemistry-based triggers, triggers related to varying points of the cell cycle, interactions of other genes which end up providing a mechanism for initiating DNA repair...the question becomes, if one has to start messing around with the redox chemistry within the cell, or mucking around with the various biochemical events in the cell cycle, you might set off DNA repair at a higher rate, but at what cost to the rest of the cell?

Direct repair methods reverse specific chemical (oxidative) modifications, they're not general ways of cleaning up DNA. Photolyase reverses the production of cyclobutane pyrimidine dimers. O6-alkylguanine alkyltransferase reverses the alkylation of guanine. They are not more general methods like the various types of excision repair pathways.

There might be hints or even partial answers to potential questions to your answers, but they're probably strewn about in the research literature, alluded to but not really strongly asserted since it's too early to say. I did get curious about the idea of transplanting a foreign photolyase into mice - Google "mouse foreign photolyase" one of these days, and you'll find the paper which describes an experiment that did just that. What I've presented is pretty much the textbook knowledge of DNA repair (as that is all I am actually familiar with myself). It would make for an interesting literature review or even meta-analysis, though.

 

[bioquest]

it's only the enzymes, nothing else that repairs DNA right

But when combined do the direct reversal methods reverse every type of oxidative dna damage? (When combined) or do they not cover all the types?

If you overexpressed all the dna repair enzymes, and that caused a higher amount of incorrectly fixed mismatched bases, would the dna repair enzymes also eventually correct the mismatched bases? Or would the incorrectly fixed mismatched bases still be a big issues

 

[Mike H]

Well, it depends on the mechanism as to what does the work of DNA repair. For something like photolyase, it's just the photolyase as I recall - it breaks the bonds between the two pyrimidine bases, and that's pretty much all there is to say. But if you look at the nucleotide excision repair pathway (see http://asajj.roswellpark.org/huberman/DNA_Repair/ner.html" ) makes note that you need the appropriate DNA N-glycosylase, followed by an AP endonuclease, followed by DNA polymerase to make the entire repair.

My understanding is that the direct reversal methods are not able to reverse every type of oxidative damage. To quote the Roswell Park website,

The huge variety of DNA-reactive chemicals in our environment combined with the huge variety of alterations that can be produced by radiation and by oxidative and free radical attack on DNA can generate so many types of damage that coping with all types of damage by evolutionary development of damage-specific DNA glycosylases would be difficult if not impossible.

If that's being said about the base excision repair pathway (which uses DNA glycosylases), I would think it applies even more strongly in the case of direct DNA repair.

As for your last two questions, not sure. Sounds like it would make for a good set of experiments, though, if they haven't already been done.

 

[bioquest]

Could/Would the increase of problems with mismatched bases etc caused by the enzymes be permanent- would they only be able to fix those errors after 2 generations/a limited amount of time or could they fix them anytime?

is that known? My friend said they only repair mismatched bases after 2 cell generations or she said something like that indicating that after an amount of time has passed, they don't correct the mistake anymore, but maybe if all the enzymes were overexpressed they would eventually fix all the mistakes?

 

[Mike H]

I am not familiar enough with the DNA repair area to say anything about such questions. I even checked my old biochem text since I'm doing some book reorganization, no such luck. These questions appear to be a step or two above, at the very least, the default undergraduate treatment.

If you could get a citation or two out of your friend regarding these phenomena she describes, it would be very helpful in either answering your question or aiding you in interpreting the articles/sources. We might even get a journal club-esque conversation going on about this topic, which could be interesting. But I, at least, am not familiar enough with this area to go ahead without some citations/references.

 

[Ygggdrasil]

Another issue to consider. It seems the cell has taken advantage of the effects of oxidative DNA damage and the repair pathways to regulate the expression of genes. Therefore, overexpression of DNA repair genes, as with any genetic manipulation, could have unintended consequences.

See also:
http://blogs.nature.com/thescepticalchymist/2008/02/chemiotics_we_had_to_destroy_t.html

 

[bioquest]

I was wondering if the radiation resistant abilities of bacteria, insects etc might lead to the solution/somewhat solution to oxidative dna damage in a mammal (I read that it's the oxidation resistance of the (dna repair?) protein(s) in Deinococcus radiodurans that help it to survive radiation, I don't know if that's because the bacteria has a high level of antioxidents in it or for some other reason, but maybe the radiation resistant protein could help with oxidative dna damage, if for example it's not resistant to oxidation just due to antioxidents?)

Then I read this though, this quote is from here http://www.madsci.org/posts/archives/2003-12/1072227809.Zo.r.html they also give past speculation on reasons for the radiation resistance and some information on why some of the reasons people thought the radiation resistance existed is wrong and list some general things that aren't behind the radiation resistance

"it is clear that insects are resistant to ionizing radiation and that this resistance is an inherent property of their cells. But it is not clear exactly what the basis of this cellular resistance is, although the dominant theory is that it relates to the relatively small amount of DNA in insect cells" would this go for bacteria and other things as well, do you think then that the radiation resistance of things and/or the radiation resistant protein in Deinococcus radiodurans would lead to a solution/somewhat solution to the oxidative dna damage problem? Why/why not?"

Yah I think people would definitely be interested in dna damage and repair to have discussions, to join a club on it or something. Um, they'd probably be way more caught up on it than I am though

I also thought this was interesting

quote from http://www.madsci.org/posts/archives/2003-12/1072227809.Zo.r.html
"The radiation resistance is inherent to the cells, since cells derived from insects are also radiation resistant when grown in cell culture. For example, a dose of 60 Gy is required to produce a 80% kill of insect cells, while doses of 1-2 Gy are sufficient to generate this level of killing in mammalian cells"

does that maybe mean it's not the antioxidents alone that are responsible for radiation resistance and that that could help with solving the oxidative dna damage issue somewhat? What do you think is responsible for the radiation resistance based on the above paragraph? thanks

Also how much would the body destroying any cancer cells that arose, if the body had that ability, help with the dna damage problem?

 

[Mike H]

Ygggdrasil - Thanks for that link, it made for very interesting reading.

bioquest - First, take a look at these three links. http://en.wikipedia.org/wiki/List_of_number_of_chromosomes_of_various_organisms and http://en.wikipedia.org/wiki/Deinococcus_radiodurans and http://en.wikipedia.org/wiki/Chromosome Read them? No? Read them first. Do it. Done yet? Good, now we can proceed.

Let me talk about the insect thing first. If you look at the list of chromosomes of various organisms, you'll see for yourself that everyone's favorite insect model organism, Drosophila melanogaster (the common fruit fly), only has 8 chromosomes, which is, as I recall, indicative of most insect genomes (barring such things like insect polyploidy). The human genome contains 46 chromosomes, which is more than five times as large as the Drosophila genome. That means humans have to run DNA repair on a much larger genome than do insects. At the site you posted, it is noted that

There was speculation in the 1950's that the number of chromosomes influenced radiosensitivity, and that insects had fewer chromosomes. There may be some truth in this speculation, as we now know that (in general) the less DNA a cell has the more resistant it is to ionizing radiation. Since DNA is the critical target for cell killing (in mammalian as well as insect cells), it is logical that the less DNA there is, the harder it is to hit. While the density of ionizations produced by X-rays is sufficient that animal size could not explain resistance, the density of ionizations is low enough that total cellular DNA content could explain resistance... So it is clear that insects are resistant to ionizing radiation and that this resistance is an inherent property of their cells. But it is not clear exactly what the basis of this cellular resistance is, although the dominant theory is that it relates to the relatively small amount of DNA in insect cells.

The take-home message from this, in my reading of it, is that the enhanced ability of insect cells to resist radiation damage is most likely due to its smaller genome. It has a lot less DNA to repair compared to mammals. To give a non-scientific analogy, think of mowing and landscaping a yard in the summer when you've got plenty of rain, so it needs to be done regularly. You're going to do a neater and better job if it's only one acre versus tending a huge 10 acre area. You're going to miss more spots, more poorly done edges, show more signs of fatigue with a 10 acre area. It has nothing to specifically do with particular aspects of insect biochemistry/DNA repair - it just means that there are fewer genes for radiation to effect in insects (8 chromosomes' worth or so versus 30 or more in various mammals) and that the insect DNA repair mechanisms have a lot less ground to cover than a typical mammalian system. It's a matter of scale, not necessarily antioxidants or specialized DNA repair mechanisms.

With regard to D. radiodurans, it has two chromosomes (chromosomes are probably not the best term to use with respect to bacteria, but that's a semantic issue I'd prefer not to get into at the moment, thank you very much). What little I know I about D. radiodurans' resistance to radioactivity is summed up in the Wikipedia article. I'm not an expert with D. radiodurans (never worked with it, only had a few mentions of it in a few classes long ago, and read the occasional news snippet about it since then), but again, you're dealing with a very small genome, there are a few copies of it in each individual D. radiodurans bacterium, and its likely relevant ability to endure very dry conditions. Here, sure, you apparently have accelerated DNA repair mechanisms (not least due to multiple copies of its genome) and high concentrations of manganese which supposedly serves as an antioxidant. I'm not sure if trying to reengineer the human species to have a couple of copies of its genome is advisable. At the very least, you'd have one heck of a time getting it past a review board. If you suggest that maybe there's a way to increase manganese intake/absorption in humans to provide a similar benefit, I would suggest you read up on the nutritional biochemistry of manganese. It is a necessary trace element, to serve as a cofactor in a number of metalloproteins, but too much can lead to undesirable side effects.

I would be sure to read the link that Ygggdrasil so kindly provided. It may be that efforts to improve one's health may turn out to be counterproductive, as there might be processes at work that one can't easily fathom.

Also, I think you might find the NIH's free online bookshelf here - http://www.ncbi.nlm.nih.gov/sites/entrez?db=books - to be an interesting and useful resource. Biochemistry, molecular and cell biology, genetics, cancer research, and more, all easily searchable.

 

[bioquest]

Is the DNA repair protein in the bacteria resistant to radiation though, (i read that it was I can look online for more info) or would that just be because of the antioxidents in the bacteria?

Also it would be impossible to use antioxidents in a way where they were never free radicals in the human body right? (assuminng free radicals they wouldn't be necessary for a human to survive although someone said to me they would be, I'm not sure)

 

[Mike H]

From the Wikipedia article on D. radiodurans in response to your first question:

Michael Daly has suggested that the bacterium uses manganese as an antioxidant to protect itself against radiation damage.[10] In 2008 his team showed that high intracellular levels of manganese(II) in D. radiodurans protect proteins from being oxidized by radiation, and proposed the idea that "protein, rather than DNA, is the principal target of the biological action of [ionizing radiation] in sensitive bacteria, and extreme resistance in Mn-accumulating bacteria is based on protein protection".[11]

It's an open question, but this hypothesis is not an unreasonable one. It could be that in D. radiodurans the presence of manganese(II) serves as antioxidant protection for the proteins. More research is needed.

I'm not sure what you're trying to get at with your second question. I can say the following - free radicals are essential in certain roles, so they are necessary, at least based on our current understanding of them. In the link that Ygggdrasil posted, antioxidant use, at least in one case, led to an increased incidence of lung cancer. It's not that they necessarily acted as free radicals, but perhaps prevented necessary free radicals from doing their jobs. (I haven't read that article which is linked there, and I'm probably not going to, given the backlog of papers I should be reading at the moment.)

 

[bioquest]

If the DNA repair protein in the bacteria is radiation-resistant but not because of the antioxidents, how could that help with dna repair in a mammal? Is it possible that all we need for good dna repair is something like oxidative/radiation resistant repair proteins/enzymes

 

[bioquest]

k, never mind, I just read what you posted I misread it a bit before sorry that was really stupid of me, I was kind of sleepy last night

But I think it would be cool to have a club or journal club-esque conversation or something where people discuss dna damage and repair, I mean I think it's a relevant, interesting, important subject to find out about and discuss and I think a lot of people would be interested in it. Since for one thing I don't know how you'd extend human life without solving the dna damage issue, unless you could regenerate whatever you needed to in the body using outside stem cells, (or using something like nanotechnology or I don't know) like embryonic ones or cells from epilepsy patients if that thing I read regarding the mcknight brain institute was correct. I don't see any real solution to the dna damage issue other than overexpressing the enzymes so I'm really interested in that, if anyone wants to talk with that about me. At first I was like, I'm going to make it my goal to find out all the unknown stuff about genes and proteins (obviously not by myself)...and then I narrowed it down to what I'm interested in which is overexpressing dna repair enzymes and now I'm interested in just that, but I'd still like to learn about other things and about genes and proteins

 

[bioquest]

My last question on it though is, are there any theoretical way(s) that could be used now that would make it so that there would be no free radicals in the brain/any specific organ that existed long enough to do damage, that could be used in a way the mammal/brain would survive?

Also could the ability to regenerate as fast as biologically possible theoretically solve the oxidative dna damage or would cells still get oxidative dna damage?

 

[Mike H]

My last question on it though is, are there any theoretical way(s) that could be used now that would make it so that there would be no free radicals in the brain/any specific organ that existed long enough to do damage, that could be used in a way the mammal/brain would survive?

Also could the ability to regenerate as fast as biologically possible theoretically solve the oxidative dna damage or would cells still get oxidative dna damage?

Short answer: I don't know.

Long answer: Asking what is or isn't theoretically possible starts to bring this discussion into ground bordering if not infringing on the overly speculative, especially when at least one of its participants (me) has not a whit of actual, useful experience in working with either eukyarotic systems or DNA repair mechanisms. If there is someone with more knowledge, who can make an educated guess based on detailed knowledge and actual experience, that's a different story. The thing to remember is that someone could follow up on an experimental anomaly this week in a lab somewhere, publish the results late next year when they've followed through, and 10 years from now we all look back and say, "Wow, that discovery really jumped us way ahead of where we thought we'd be at the moment." Or they could not, and people miss some observations or make some understandable but ultimately incorrect conclusions, and suddenly we're far behind the curve as it was originally imagined.

I don't mean to discourage you from asking questions, but these are questions which demand a far more extensive knowledge of the field in question and its literature than is typically passed along to non-specialists in the field, at least from what I can tell.

I would think about perhaps going back, really delve into the links that have been shared in this thread (and the citations/references contained within those links), and work your way through things in a more methodical and less rushed manner. Right now I feel like you're asking us to run up to the peak of Mount Everest ("It's only five and a half miles, everyone, it's not that far of a distance!") with you, instead of making our way slowly through the various base camps at different altitudes. Read up on DNA repair. Read up on what triggers/initiates DNA repair. Read up on antioxidant mechanisms, on the roles that free radicals do carry out in biological systems. Read up on alternate hypotheses on how to extend an organism's life like caloric restriction. (No, I have no information on it beyond the Wikipedia entry. Sorry.) Read up on how scientists actually overexpress or silence gene expression in cells. Take notes, draw yourself diagrams, sketch out pathways, really take your time and go step by step.

 

[bioquest]

Are there synthetic DNA repair enzymes that are being used/made or anything that could solve the oxidative dna damage issue? I mean I know they've made synthetic DNA

 

[Mike H]

Are you talking about known DNA repair proteins that were the focus of site-directed mutagenesis studies to modify their behavior or proteins designed from scratch? If the former, I have not a clue. I imagine that literature would be quite an effort to map and analyze. If the latter, there are only a few published results of proteins being designed from scratch to do actual chemistry (not just ligand or cofactor binding) and none of them, as I recall, had anything to do with DNA repair.

You should probably look up "protein engineering" and "protein design"/"de novo protein design" for a better idea about the latter, if that's what you were asking about in your question. It's a very interesting but very challenging field.

 

[bioquest]

are there 2000 year old trees or anything, or is it just the bark or something from those that are alive after thousands of years? I thought there was a type of tree that lived to 2000

 

[bioquest]

I mean, is it just the bark on trees like this http://www.Earth'sky.org/teachers/article/giant-sequoia that live a really long time, or is it the tree itself, like could the dna damage ability etc of these trees be transferred to animals and could the animal then live a really long time, or it is just the bark or something on the trees that live a really long time?

it says here
The oldest big trees are about 3000 years old.

http://www.icogitate.com/~tree/species/sequoiadendron.giganteum.htm

 

[bioquest]

Flooding cells with antioxidents/things that kill free radicals wouldn't stop all oxidative dna damage because the mitochondria/free radical production and the mitochondrial dna are so close together right?

Also sorry if this is an ocd question but you wouldn't be able to change a human into a different organism than a human using genetic manipulation or something right?