Monday, January 16, 2023

Transformer p245 and onwards: glutathione

In Transformer Nick Lane has an interesting discussion of the roles of glutathione in redox chemistry. He seems to have been spurred to look at this when his research group had a WTF moment with fruit flies. They've not published the work yet so Transformer is the only place to read about it.

They had highly, highly inbred (identical) fruitflies with respect to nuclear genes. They maintained them as identical as possible. Different strains had mitochondria with minor variations in mtDNA, within normal limits, but with differences of "fit" to the (all identical) nuclear genome. Some combinations gave better ETC characteristics than others.

To see if the variations in function were due to ROS flux they treated the flies with n-acetyl-cysteine, a glutathione precursor, producing an excellent scavenger of ROS. You know:

2G-SH  +  H2O2  ->  G-S-S-G  + 2H2O

and all should be well, or at least better.

That didn't work out too well. Males were more or less OK. Most females were "seedy" on NAC. In one strain all of the females, only, died. They checked glutathione levels and the NAC appeared to be doing what it was supposed to do, lots of glutathione.

Lane's idea, which is rather insightful as regards life, is that a body will tolerate an ROS flux within certain limits. To limit excessive ROS formation cells are willing to limit oxidative phosphorylation by deactivating complex I. Compromising ATP production is considered acceptable in order to limit ROS generation to "tolerable" limits.

This is not really surprising. If we think about uncoupling proteins their core function is to dissipate delta psi to a voltage which will not generate many ROS (less than ~140mV), at the cost of decreased ox-phos.

At a guess you might be able to limit/localise the glutathiolation effect to those proteins which are responsible for excess ROS, so glutathione glutathiolates the cysteines within said protein (in this example complex I) in proportion to ROS being generated. Like this

G-SH  +  Prot-SH  +  H2O2  ->  G-S-S-Prot  +  2H2O

Glutathiolation has evolved to alter the function of a protein in such a manner as to decrease ROS generation, even if that includes a decrease in oxidative phosphorylation.

Lane's guess is that female flies, with their high demand for ATP for egg production, couldn't cope with the drop in ox-phos mediated by glutathiolation of complex I.

These flies died in order to limit ROS production.

Have I ever mentioned that ROS are central to, well, everything?


Supplementary thought: Perhaps we could better phrase it that oral NAC raises glutathione to levels in excess of those which are already ideal, so hyper-glutathiolation causes death by excess ox-phos limitation. This would be particularly problematic for flies with slightly more ROS generation than others. The flies were okay provided ROS and glutathiolation of complex I were at physiological appropriate levels.

Fascinating in view that NAC/glutathione appears to be a pretty Good Drug in general terms. But, as always, over riding evolution has costs.


  1. Wow. Super interesting!
    Thank you for sharing Peter! :)
    (Have a great day, everyone!)

  2. It's always a pleasure to stumble across new rabbit-holes although there are more of them than hours in the day. It is amazing to think of something the size of a fruitfly with its staggering complexitity. Life is so good eh?

    NAC and glutathione have one interventional use in humans, to reverse a glutathione crisis which can be caused by the toxic metabolite of paracetomol/acetominaphen. Which led me to wonder if drosophila have livers? Well no, they have a weird circulatory setup with no organs per se so panadol might not help them recover from a glutathione overdose.

    But, heading off on a tangent they have fascinating lipid metabolism via liver-like cells. (Tldr - its controlled by insulin!)

    "Fly Foie Gras: Modeling Fatty Liver in Drosophila

    ( ref snip) .... further extends the known functional homology between vertebrates and flies in this process. Specifically, they uncover a novel hepatocyte-like role for a previously mysterious class of specialized cells in Drosophila larvae called oenocytes

    Flies store lipids in the form of droplets in the fat body (FB), a specialized tissue that resembles vertebrate fat in many respects. In response to starvation, a sensor mechanism in FB cells that is controlled by insulin/IGF (IIS) and TOR signaling triggers the aggregation of lipid droplets into larger structures, the upregulation of lipases such as the ATGL-like enzyme Brummer, and the release of lipids into the hemolymph (Colombani et al., 2003, Gronke et al., 2005). How these lipid stores are processed for further use by larval tissues has long been a mystery but is now solved by the work of Gutierrez et al. (2007). Using classic oil red O staining, the authors first show that larval oenocytes, which are visible in well-fed animals as segmentally arranged clusters of large cells that are largely devoid of fat stores (Gould et al., 2001), accumulate vast quantities of lipid droplets under starvation conditions "

  3. i've read about good uses for NAC, but it seems to be a double-edged sword, so wield carefully!

    e.g. Impact of N-acetyl-l-cysteine on SARS-CoV-2 pneumonia and its sequelae: results from a large cohort study

    "133 patients died during hospital stay (91 in the NAC group and 42 in the control group)"

    My guess is the ARDS (i.e. cascade of linoleic acid oxidation) that occurs in covid-19 was overwhelmingly suppressed by NAC to the point where even the *useful* inflammatory responses to fight viruses were inhibited

  4. Wow Pass, nice insight into fruit flies. I wonder how they treat their *natural* hangovers after drinking excess fermented fruit juice..... Surely not paracetamol?

    raphi, next up might be SkQ1 to ameliorate LA lipid oxidative crap. But perhaps this comes from the Wrong Place. After all, if we're going to ban Tchaikovsky performances on the basis of nationality how could we allow SkQ1? Happily (???sadly???) I doubt it would work any better than NAC.


  5. 'Fascinating in view that NAC/glutathione appears to be a pretty Good Drug in general terms. But, as always, over riding evolution has costs.'

    NAC is a popular supplement. Ever since Peter started pointing out the importance of ROS for signaling, I've been wondering about the modern fixation on antioxidants.

    This is timely as I'm just finishing the Sam Apple book Ravenous, about Otto Warburg, the Nazis, and cancer; it's light on scientific detail but the historical context is very interesting. Gary Taubes pointed out in Good Calories, Bad Calories that the Germans had made great strides in metabolic science which were subsequently obscured by postwar attitudes toward nazi evildoing.

    After reading this post, I thought I finally understood that a cell could fall back on glycolysis if the mitochondria were damaged (which Warburg insisted, to the end, is the cause of cancer, mitochondrial dysfunction). I do know that glycolysis is not nearly as efficient in producing ATP as is oxidative phosphorylation, although everything I've read about the Warburg Effect fails to explain why it's such a big deal that glycolysis takes place in the absence of oxygen.

    Here's a series of animations covering glycolysis to ox phos, which mostly served to confuse me further

  6. Apologies for being off-topic, but passthecream's remark abou the complexity of life provides an excuse to offer this day-brightener.

  7. Fruitflies are more likely to cause fermenting fruit than be caused by it, but perhaps they have tiny little Beroccas to help with hangover? In reality ethanol is probably as good a source of acetate for them as it is in humans ( etoh -> acetaldehyde then to acetate via ALDH) also cyp2e1 and catalase are involved. It seems as though humans have picked up a variety of ways of handling it as a food source over long evolutionary history from vinegar flies to where we are now.

    --- problems arise if there is too much aldehyde generated or if methanol is involved. Methanol is oxidised via adh to formaldehyde, very nasty.

  8. I've been following GlyNac research. GlyNac = supplementation of two glutathione precursors, NAC and glycine. GlyNac research has shown only positive benefits by restoring glutathione levels in aged organisms to the level of young organisms.

    So why did these female fruit flies die? Did their levels become higher than that of healthy young animals?

    Researchers in one of the studies said that the GlyNac supplementation did not cause reductive stress:
    "As discussed above, cells need to combat OxS and simultaneously avoid reductive stress. As seen in this report, the heart, liver and the kidneys have differing GSH concentrations. Indeed, every organ of the body maintains its own GSH concentrations based on their metabolic activity, generation of ROS and cellular needs. Exogenous compounds with antioxidant potential run into the problem of dosing—too much can induce reductive stress, and too little will not combat OxS. This is further complicated by the fact that every organ maintains a different concentration of GSH, and exogenous antioxidant dosing is unlikely to correct GSH deficiency and OxS, while simultaneously avoiding reductive stress in all organs. GlyNAC does not attempt to override or replace cellular defenses to combat OxS and reductive stress. Instead, by providing precursors to boost GSH synthesis and concentration, GlyNAC works by supporting the ability of cells to autoregulate their own GSH—thus GlyNAC can correct GSH deficiency and OxS in every cell and organ in the body, without inducing reductive stress."

    "GlyNAC supplementation did not cause an overshoot of GSH concentrations above levels seen in YA, and did not lower markers of OxS below levels in YA, suggesting that GlyNAC supplementation allows cells to successfully autoregulate cellular GSH concentrations required to combat OxS, while simultaneously avoiding reductive stress"

  9. @Cave. Help me out a bit. Be gently, please! Isn't the issue with cancer that the Krebs cycle flips and goes backwards pumping out sugars? I thought I'd got the hang of most mitochondrial issues but learning that gobsmacked me a bit - although I was aware of Warburg's fermentation ideas. Anyway, I'm going to buy that Warburg book. Not being too heavy on detail will suit me just fine!

  10. Captain Sunset: "Isn't the issue with cancer that the Krebs cycle flips and goes backwards pumping out sugars?" Thanks to you asking, I learned a lot this morning. There actually is a reverse Krebs cycle. "The reverse Krebs cycle is proposed to be a major role in the pathophysiology of melanoma. Melanoma tumors are known to alter normal metabolic pathways in order to utilize waste products. These metabolic adaptations help the tumor adapt to its metabolic needs. The most well know adaptation is the Warburg effect where tumors increase their uptake and utilization of glucose."

    At least now I know why glycolysis in the absence of oxygen is a Thing, having just now discovered that the definition of fermentation is different from the commonly understood meaning. "Fermentation is a metabolic process that produces chemical changes in organic substrates through the action of enzymes. In biochemistry, it is narrowly defined as the extraction of energy from carbohydrates in the absence of oxygen." (wikipedia)

    The book 'Ravenous' is mainly historical. Cancer was becoming alarmingly widespread in the 1900s; the Germans were avidly pursuing metabolic and environmental causes of cancer, including the Nazis. Auschwitz had a thriving organic garden tended by inmates. Hitler, whose mother died of breast cancer when he was a teenager, was a wacko vegetarian. This research was largely forgotten thanks to World War II, and then the world became fixated by genetics with the discovery of DNA structure in 1953. More general interest in the metabolic aspects of cancer revived in the 1990s. On finishing the book I learned in the acknowledgments that Gary Taubes had a not-insignificant influence on the writing. Incidentally, Krebs was a student of Warburg. Warburg was a huge deal during his time, and you'll recognize a lot of the people he associated with, including Einstein.

    Travis Christofferson's booik 'Tripping over the Truth' is supposed to be a layman's summary of Thomas Seyfried's 'Cancer as a Metabolic Disease'. I've read the first but not the second; this interview looks interesting

    Having recently gone through a cancer blip myself at a major cancer center, I can tell you the mainstream is still fixated on genetics and has absolutely no interest in metabolism or diet, even as an adjunct treatment, at least where I went; I made it a point to try to talk to every single person I interacted with. There is some research being done at other large cancer centers, which is encouraging.

  11. Cap'n; Newt; this Peter blog from 2019 broadens the picture somewhat (and the -2 blog on the same topic)

    “It is worth noting that the ability to reduce oxygen at the cell surface to support glycolytic metabolism may be a fingerprint of the early endosymbiotic origin of eukaryotic cells”

  12. you're going to like this Peter "Ants Live 10 Times Longer by Altering Their Insulin Responses"

  13. Pass, thanks. To ask what is probably a dumb question, per Peter's 2019 blog post "Cell surface oxygen consumption (2)" at

    ...which concludes "In rho zero anabolic cancer cells cell surface oxygen consumption can be as much as 90% of the total oxygen consumption of the parent cell line" (rho zero cells have no ETC)...

    I'm trying to figure out how this squares with the definition of glycolysis (fermentation) which is "In biochemistry, it is narrowly defined as the extraction of energy from carbohydrates in the absence of oxygen." Does the definition not matter because the oxygen is consumed at the cell surface and only serves to create NAD+? IOW the oxygen never actually penetrates the cytosol?

  14. Cave & Pass. Tks! I've got homework to do... Algae in aquaculture is a big thing, and that used to be just about where I stopped with the science, but it does get much more interesting.

  15. 'Newt, I'll have to catch up on it myself :)
    I'm falling behind on my rabbit hole quotient atm.

    But the group whose work that is moved the scene of action more to to the plasma membrane than the mitochondria, and there are some similar functional structures there ( on the PM versus the MM).

    To further complicate matters take a look at Michael Levin's work. Consider his frog derived xenobots and their organising/recruiting capabilities as throwing a different light on cancer cells. It's not his main area of interest but mind blowing nonetheless!

    (Link thanks to Peter).

  16. Btw, pick your plenarian - they reproduce sexually and asexually, and

    "... planarians reproduce by ripping themselves into a head and a tail piece, which subsequently regenerate into two new worms"


  17. My grandfather was the head naturalist for Cook County (Chicago) and he used to send us his monthly newsletter when I was a child. One was about earthworms and I remember it vividly: "If you cut a worm in half, each half grows a new tail and then the tail/tail half starves to death."

    Sounds like planarians are a little more sophisticated.

  18. If you split the head end of a planarian in half sideways each half head regenerates into a whole new head so you get a two headed worm. Supposedly both heads have the same ' memories'. Zaphod Beebleworm?

  19. Zaphod indeed. That is totally awesome.



  21. Off topic I know, but the following is very good re the bloody SARS2 Spike and the brain, plus a wee bit more.

  22. Sorry, meant to say watch from 19 mins in.


  23. @Pass "" Yep, all very, very intriguing. Thankfully, cancer hasn't personally passed my way as yet, but the whole O2/cell interaction is indeed fascinating. All too clever for me, but I would be interested to know if any here had strong feelings as to pointers of such. It frustrates me that good out-of-box thinking with cancer is a bit verboten. I have to say I find a lot of what comes up on the subject (outside of places like this) is utter crap. It 'smells' rotten.

  24. Hi Cap'n

    I had a little cancer blip a couple of years ago so I read some about it. Metabolic theories of cancer are more involved with glucose than oxygen; Thomas Seyfried might be a good place to start. I understand his book Cancer as a Metabolic Disease is kind of technical, but a more populist overview would be Travis Christofferson's Tripping Over The Truth. There's a lot about the Warburg effect in there.

    Given the current cultural antipathy toward ROS and popularity of anti-inflammatories, I would be interested to read things to the contrary also.

  25. @Pass 'n Cave, Tks! I'll go with the Seyfried book. I think I can handle it. And, yes, I'm with you on the glucose & ROS. Both are what really got me delving in matters. That, and of course Nick Lane's books.