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how statins work 3: the beginnings of cholesterol, from Acetyl-CoA

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Coenzyme A – an acetyl group attaches to the -SH shown in red

So how is cholesterol made in the body? We need to know this in order to understand how statins inhibit or interfere with this process.

I’ve shown the actual structure of cholesterol in part 1 of this series, but remember it’s a sterol, which is a steroid – four carbon rings with hydrogen atoms attached – in which one of the hydrogens is replaced by an alcohol group. The particular form of sterol called cholesterol, with a 7-carbon chain attached to the end-carbon ring (the D ring), and three methyl groups attached to specific carbons in the rings and chain (it’s better to look at the skeletal structure in part 1). There are precisely 27 carbon atoms specifically placed within the molecule.

I’m using a set of videos to understand how cholesterol is synthesised – it might be best to look at them yourself, but I’m writing it all down to improve my own understanding. So we start by understanding something about acids and their conjugate bases. Apparently an acid is a molecule which is capable of donating protons into solution. Take pyruvic acid and its conjugate base pyruvate. Here’s what Wikipedia says about them:

Pyruvic acid (CH3COCOOH) is the simplest of the alpha-keto acids, with a carboxylic acid and a ketone functional group. Pyruvate, the conjugate base, CH3COCOO, is a key intermediate in several metabolic pathways throughout the cell.

I don’t understand the first sentence, but no matter, pyruvic acid is a 3-carbon molecule with a carboxylic acid at one end and a ketone group in the middle of the molecule (according to Britannica, a ketone is ‘any of a class of organic compounds characterized by the presence of a carbonyl group in which the carbon atom is covalently bonded to an oxygen atom. The remaining two bonds are to other carbon atoms or hydrocarbon radicals)’. The proton that comes off the oxygen of the alcohol group of the pyruvic acid can be donated into the surrounding solution, increasing its acidity. The pyruvic acid is thus transformed into its negatively charged conjugate base (it’s no longer capable of donating protons but it can receive them). This is the case with all acids in the cytoplasm of cells. As inferred in the quote above, conjugate bases are vital components of biosynthetic pathways. Most of the molecules in the cytoplasm will exist as pyruvate at a physiological pH of around 7.5.

Next – and hopefully this will become clear eventually – we’re going to look at two molecules, NAD+ (nicotinamide adenine dinucleotide) and NADP+ (nicotinamide adenine dinucleotide phosphate). They transport electrons, and are capable of accepting a hydride anion, which is a hydrogen atom with a negative charge. The normal hydrogen atom, called protium, has a proton and an electron only. When it donates away its electron it becomes a hydrogen cation, and when it gains an electron it becomes a hydride anion.

NAD+ is an adenine organic base bound to a ribose sugar. Then there are two phosphate groups coming off the ribose sugar, the second of which attaches to another ribose sugar. This second ribose sugar has nicotinamide attached to it (see below),

in which the phosphate groups are magenta-coloured circles. To explain something about ribose sugars, here’s something from Pearson Education:

The 5-carbon sugars ribose and deoxyribose are important components of nucleotides, and are found in RNA and DNA, respectively. The sugars found in nucleic acids are pentose sugars; a pentose sugar has five carbon atoms. A combination of a base and a sugar is called a nucleoside. Ribose, found in RNA, is a “normal” sugar, with one oxygen atom attached to each carbon atom. Deoxyribose, found in DNA, is a modified sugar, lacking one oxygen atom (hence the name “deoxy”). This difference of one oxygen atom is important for the enzymes that recognize DNA and RNA, because it allows these two molecules to be easily distinguished inside organisms.

So, just for my own understanding, nucleotides include phosphate groups. NAD+ is a dinucleotide, with two nucleotides (ribose sugars with phosphate groups attached), attached to adenine and to nicotinamide molecules. Also, NAD+ has a positive charge around the nicotinamide – on its nitrogen atom.

NAD+ becomes neutralised by accepting a hydride anion (one proton and two electrons) and becomes NADH, or reduced NAD. Now, remembering NADP+, it has an extra phosphate group on the ribose sugar of the adenine nucleotide (also called an organic base, apparently). Like NAD+, NADP+ can accept a hydride anion (becoming reduced NADP) and then later exchange it in another reaction. Effectively these molecules are electron carriers, collecting electrons and transporting them to where they’re needed for other reactions.

Now to introduce something else completely new for me – Acetyl-CoA (acetyl coenzyme A). A quick grab again, this time from Wikipedia:

Acetyl-CoA is a molecule that participates in many biochemical reactions in protein, carbohydrate and lipid metabolism. Its main function is to deliver the acetyl group to the citric acid [Krebs] cycle to be oxidized for energy production

Acetyl-CoA is found, and presumably produced, in mitochondria, and as part of this cholesterol-synthesising pathway it needs to be removed from the ‘mitochondrial matrix’. What’s that, I ask. So here’s a bit about the mitochondrial matrix, from yet another source, this time Study.com:

The mitochondrion consists of an outer membrane, an inner membrane, and a gel-like material called the matrix. This matrix is more viscous than the cell’s cytoplasm as it contains less water. The mitochondrial matrix has several functions.It is where the citric acid cycle takes place. This is an important step in cellular respiration, which produces energy molecules called ATP. It contains the mitochondrial DNA in a structure called a nucleoid. A mitochondrion contains its own DNA and reproduces on its own schedule, apart from the host cell’s cell cycle. It contains ribosomes that produce proteins used by the mitochondrion. It contains granules of ions that appear to be involved in the ionic balance of the mitochondrion.

So basically this matrix is like (or equivalent to) the cell’s cytoplasm, only more viscous, and contains ribosomes, one or more nucleoids and ionic granules, inter alia.

Acetyl-CoA is essential to the biosynthesis of cholesterol, and is found initially in the mitochondrial matrix, and we need to look at the pathway for its removal from that matrix into the cytoplasm, where all the action occurs.

Intruding into the mitochondrial matrix from the (quite impermeable) inner cell membrane are the cristae, which give the membrane more of a surface layer for interactions. This inner membrane is the site of oxidative phosphorylation. What’s that, I ask. Well, it’s key to the production of ATP, and at least I know that ATP is the ‘energy molecule’, and that it’s produced in mitochondria. Here’s something about the process from Khan Academy:

Oxidative phosphorylation is made up of two closely connected components: the electron transport chain and chemiosmosis. In the electron transport chain, electrons are passed from one molecule to another, and energy released in these electron transfers is used to form an electrochemical gradient. In chemiosmosis, the energy stored in the gradient is used to make ATP.

So a strong proton gradient is built up across the inner membrane of the mitochondrion. It’s a concentration gradient but also an ‘electrical potential difference’ gradient, so that the electrical potential within the matrix is lower, by some 160 millivolts, than that across the inter-membrane space. The protons within this space are unable to pass back into the matrix. The only way they can get back into the matrix is by means of ATP synthase which can harness the energy from the protons as they move down the chemical and electrical gradient, and use that energy to bind ADP to inorganic phosphate to create ATP.

I don’t fully understand all that, but the main point here is that the mitochondrial inner membrane is very ‘tight’, which makes it difficult to transfer Acetyl-CoA out of the matrix and into the inter-membrane space, from which it can more easily diffuse through the more permeable outer membrane into the cytoplasm.

The structure of Acetyl-CoA: it consists of an acetic acid molecule (CH3COOH) with a thioester link to the thiol group of a coenzyme A molecule. The importance for us here is this thiol (HS) group, which is similar structurally to an alcohol (HO) group, as sulphur has similar properties to its periodic table neighbour, oxygen. So thiol groups can be linked to carboxylic acid groups as alcohol groups can. Acetyl essentially means acetic acid with the alcohol removed. To get this Acetyl-CoA out of the matrix, it is first bound to oxaloacetate, a four-carbon molecule, to create citrate, the first molecule of the citric acid cycle. This citrate can be passed through the mitochondrial inner membrane and into the cytoplasm where it can be converted back into Acetyl-CoA.

So the conjugate base, oxaloacetate, has carboxylic acid groups, attached to the first and fourth carbon atoms, that have lost their protons into solution. An enzyme within the matrix is able to combine oxaloacetate with Acetyl-CoA and water to create citrate…

Okay, this is proving to be a much longer story than I might’ve hoped, but I like to be thorough – and in reality I’m still not being thorough enough. There’s a lot of rubbish on the internet about statins, much of it self-serving in one way or another, so I’ll just keep plodding along until I feel at least halfway informed about the matter. Meanwhile, you just keep getting on with your work, and don’t mind me.

References

Cholesterol biosynthesis part 1, by Ben1994, 2015

Cholesterol biosynthesis part 2, by Ben 1994, 2015

https://www.britannica.com/science/ketone

http://www.phschool.com/science/biology_place/biocoach/bioprop/ribose.html

https://en.wikipedia.org/wiki/Acetyl-CoA

https://study.com/academy/lesson/mitochondrial-matrix-definition-function-quiz.html

https://www.khanacademy.org/science/biology/cellular-respiration-and-fermentation/oxidative-phosphorylation/a/oxidative-phosphorylation-etc

Written by stewart henderson

October 14, 2019 at 5:26 pm

HIT, mitochondria and health

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comparing-hit-with-other-exercise

 

and for a gentler form of exercise...

and for a gentler form of exercise…

Jacinta: Well now, I know you’re dying to explore the recently touted benefits of your favourite exercise, so let’s have it.

Canto: Yes, I’m very much a HIT man, that’s high intensity interval training, highly recommendable because it takes so little time and only requires an exercise bike. I was put onto it by one of Michael Mosley’s documentaries, though I’ve been a rather theoretical enthusiast in recent times, having trouble overcoming my laziness and my pain-avoidance tendencies, because though it’s short exercise it is a little painful.

Jacinta: So the recent Catalyst episode has brought your enthusiasm surging back?

Canto: Naturellement, especially as it brings with it some new research to focus on. Mitochondria – what do you know about them?

Jacinta: That they are organelles in our cells, believed to have originated as bacteria but to have united with our eukaryotic cells way back in time in a process known as endosymbiosis. They’re also responsible for producing ATP, the energy molecules… though I’ve no idea how, or what an energy molecule actually is.

Canto: That’s music to my ears.

Jacinta: The dulcet tones of ignorance?

Canto: In the country of the blind the one-eyed science pundit is king, and I’d rather be a king than a commoner, so hear ye, my subject.

Jacinta: I may be blind but I’m all ears, Your Majesty.

Canto: Well, as the Catalyst program tells us, mitochondria are about a billion times smaller than a grain of sand, but the world at nanoscales has really opened up to us in recent decades. Mitochondria are good for us, and the more the merrier. And the evidence is that HIT exercise can not only increase the production of mitochondria but increase their function.

Jacinta: So how do we produce mitochondria?

Canto: Are you going to keep interrupting me with questions? Okay, the production of mitochondria relies on our oxygen intake. The story goes that we fill our lungs with oxygen and it enters the bloodstream for a specific purpose…

Jacinta: Hang on, we fill our lungs with air, not just oxygen, so how does the oxygen get separated, and how does the blood take up the oxygen? Aren’t you skipping a few steps here?

Canto: Yes, go and research it yourself and you can report on it next time. The destination of this inhaled oxygen is the mitochondria. There are billions of these mitochondria in our musculature, though the more fit and trained up you are, the more you’re likely to have. Mitochondria apparently comprise some 10% of our body mass, which I’m sure will come as a surprise. Now oxygen, as you know, acts as a corrosive through the process known as oxidation, which involves the loss of electrons…

Jacinta: Hang on…

Canto: Please shut up. So oxygen can have a negative effect on proteins, enzymes and even our DNA, but mitochondria uses this corrosive electron-stripping power to break down nutrients and to create energy in the form of adenosine triphosphate (ATP). Don’t ask! Of course this doesn’t just happen in humans but in all other mammals and complex creatures, and in plants. And that brings us to physical fitness, and the VO2 Max, which is, essentially, the measure of the fitness of our mitochondria. The term stands for volume (V), oxygen (O2), and of course maximum, though generally those concerned with aerobic fitness don’t make the association with mitochondria, they’re just looking at increasing their maximum oxygen consumption levels. Now it’s not an easy thing for impoverished nonentities like us to find out what our VO2 Max is, but it’s probably pretty pathetic. It’s something that endurance athletes tend to obsess about as they try to improve their performance – I believe rowers in particular have some of the highest levels. I notice there’s at least one VO2 Max app on the market – going very cheap too – but I’d be very sceptical about its reliability. In the testing facility shown on Catalyst they measure it via a version of HIT. They get the subject to ride an exercise bike, building up speed till she’s going as fast as she can, and she can go no faster and starts slowing down. That peak represents her VO2 Max. She will be tested 16 weeks later, after a mere 6 minutes of HIT a week, and you can bet your rented house that her VO2 Max will have substantially improved.

Jacinta: So for us low-lifes – excuse my interruption – who can’t easily or cheaply measure improvements in our VO2 Max or, say, our fat to muscle ratio, we just have to feel the difference in aerobic fitness, mitochondrial health and the like…

Canto: Yeah, and your weight will go down too, if you’re carrying a bit extra, as we both are. And the exertion will make you feel better and healthier, I guarantee it. We all know that the placebo effect is real after all. But seriously, I’m sure if we keep to a regime of HIT – say 3 bursts of 20-second full-pelt pedalling interspersed with a minute or so of more relaxed pedalling, or even if we start with 10-second bursts and then 15-second bursts, maybe eventually getting up to 30-second bursts, we’ll feel it getting easier, and it won’t be purely subjective even if we have no way of objectively measuring it.

Jacinta: But shouldn’t we consult a doctor beforehand? I already feel a heart-attack coming on.

Canto: I know you’re joking, but certainly anyone who has any kind of heart condition, or are diabetic or pre-diabetic or have any other serious chronic condition should discuss it with their GP, but really, apart from your couch potato tendencies, there’s nothing wrong with you.

Jacinta: You’re right, and I’m looking forward to the challenge, even though I’m already a to-die-for, effortlessly slim, perpetually twenty-two year old intellectual beauty..

Canto: And I’m the ultimate metrosexual hipster of indeterminate age and shoe size, discreetly tattooed and tucked…

Jacinta: Ah, yuck, you stupid twat, tattoos are the most repugnant fashion development of all time. At least you’re not a spornosexual, yuk, stay away from the gym or I’ll never speak to you again .

Canto: Promise? Anyway, around 35 is the average VO2 Max, but that’s a bit meaningless for us low-lifes as you say. Top athletes have levels in the 60s and 70s, with the highest ever recorded being around 96 or 97 for humans, but some mammals – like racehorses and Siberian sled dogs – can reach much higher levels. But there’s also going to be a big improvement in your fat-to-muscle ratio with regular bouts of HIT. In the Catalyst episode, the reporter took a DEXA body composition scan to measure this ratio. It also measures bone density. DEXA stands for Dual Energy X-ray Absorptiometry, that means you’re subjected to 10 minutes of very low-dose x-radiation at two different energy levels. It measures the relative densities of the different tissues. You can get this scan done in Adelaide, for a baseline measure, but it’ll probably cost an arm and a leg.

Jacinta: One way to lose weight. Cheaper to just take it for granted that you’re getting more muscular with every HIT.

Canto: Spoken like a true scientist. But generally, inactivity itself is a health problem, and anything that raises your metabolism, as HIT most definitely does, will be good for you, if it doesn’t kill you. And of course one of the most exciting findings in recent times is that your VO2 Max can be raised, with all the associated health benefits, without spending crazy amounts of time and money at the gym.

Jacinta: So how did they make this discovery?

Canto: Well I suppose they were doing a lot of experimenting and testing around the health benefits of exercise, but one test, a Wingate test, involved 30 seconds of all-out pedalling on an exercise bike, repeated a few times between periods of rest, to make up to two or three minutes of full-on exercise per session.

Jacinta: And this was for already-athletic types, right?

Canto: Yes – not advisable for middle-aged or post-middle-aged couch potatoes to start on that regimen. I’m currently doing three fifteen-second bursts, building up to 20-second bursts, then up to 30 seconds and no more. So researchers found that endurance levels can be dramatically improved after just six minutes or so of this kind of exercise. A doubling of endurance capacity, no less. Compare this to the current recommendations of 150 minutes a week. Who ever does that, apart from gym junkies?

Jacinta: So, it’s like this incredible short-cut to health.

Canto: Well of course it isn’t the solution to all ills, but among other things such a quick turn-around is a great motivator towards a healthier lifestyle all round. And it doesn’t have to be an exercise bike – you can adapt it, for example you can get yourself outside and do interspersed 30-second sprints, but I hate running and I’ve got a gammy knee so I’ll stay on the bike.

Jacinta: So, have they looked more into the actual science of this? What’s happening here?

Canto: Well again it seems to be about sucking in oxygen and providing a drug hit to the mitochondria. They did this rather nasty experiment with mice, genetically modifying them so that their mitochondrial DNA wasn’t functioning properly – their mitochondria were getting worn out. They looked pretty sorry-looking compared to the control mice, prematurely ageing as evidenced in their fur, their neural activity, heart function and sensory abilities. Their life-span was about half that of normal mice, and no drugs improved the situation.  Then they set them on a treadmill regularly, 3 times a week, at a brisk pace, for 45 minutes each session, which you might think would’ve killed them off all the more quickly, but the result was a spectacular improvement in mitochondria production and overall health and energy levels.

Jacinta: And this was in genetically modified mice?

Canto: Apparently so. The program didn’t go into detail about that, except to say that the bad mitochondria were apparently being selected against. Now of course we’re talking about mice here, and this was looking at endurance fitness rather than HIT, but it’s been shown that HIT does all the right things, and in some areas performs better than endurance training. Reductions in blood pressure, improvements in insulin sensitivity, in muscle to fat ratio, in VO2 max all in a matter of weeks, but the really interesting finding was that with HIT, improvement in mitochondrial function was significant – which wasn’t the case after endurance training.

Jacinta: How do they know that?

Canto: They took muscle samples and measured the ability of the muscles to produce oxygen – basically a measure of mitochondrial function. After just four weeks of HIT, mitochondrial function improved by up to 30%, while endurance training over the same period showed little or no change.

Jacinta: Wow. Doesn’t say much for endurance training.

Canto: Well endurance training does improve your VO2 max and it’s hardly bad for you. But the thing with these quick sprints is the difference at the muscle level. Sports medicine distinguishes between fast-twitch, slow-twitch and intermediate muscle fibres. HIT uses a wider range of muscles and muscle types than endurance work, and that seems to be the key. Improvement in mitochondrial function confers a heap of benefits, so this kind of exercise wards off neurological and other conditions, including muscle weakness and epidermal deterioration, the tell-tale signs of ageing. In fact all exercise does this. Ever heard of the stratum corneum?

Jacinta: Mmmm, corneum, cornea, isn’t that part of the eye?

Canto: Excellent guess but wrong in this case. The stratum corneum is the top layer of the epidermis, the skin. It starts to thicken as you age, and the layer underneath gets thinner as your mitochondrial function reduces. You can slow down that process quite significantly with regular exercise. They did skin biopsies of sedentary people over 65 before and after endurance training. After just 3 months the skin showed great improvement – a 20 to 30 ‘youthening effect’, according to one researcher. The dead outer layer thinned, and the dermis, full of collagen fibres, thickened. So, clearly, you’re never too old to start.

Jacinta: Or never too young. So okay I’ll start.

Canto: Great, but let me describe one more impressive study, being done on menopausal women using HIT. Menopause is about a major decline in estrogen, which has serious vascular, heart and metabolic effects, as well as insulin resistance. You tend to produce a lot of bad visceral fat which negatively affects the liver, due to the over-production of cytokines – but that’s another story. Anyway, the women were given a sprint regime, of just a short period of fast peddling interspersed with more relaxing peddling, amounting to eight minutes of fast but not hard exercise all up. The results of this research haven’t been published yet, but the women’s self-reporting is all very positive, which isn’t surprising. The research is also based on previous research with obese young men, and the exercise proved very effective. Visceral fat is generally much easier to reduce than subcutaneous fat.

Jacinta: Okay, so we’re going to do this?

Canto: Absolutely. And finally, here are some links.

 

The Catalyst episode, http://www.abc.net.au/catalyst/stories/4319131.htm

http://www.tabataprotocol.com

https://newsroom.unsw.edu.au/news/health/sprint-fight-fat

High-Intensity Training and Changes in Muscle Fiber, [www.springerlink.com/content/1137px7x66667132]

Written by stewart henderson

October 16, 2015 at 8:34 am