FUNdamentals 1

09/10/08 (Dr. Baggot)

11:00 – 12:00

Introduction:

Acetyl and Malonyl

  • …linked to each one of the dimers. This is the reason why we have substrate channeling. It is handing off intermediates from one active sit to another active site. Once we get up to palmitate then it is released.

Origin of fatty acid substrates

  • Where do we get the substrate to make fatty acids? It comes from the mitochondria in the TCA cycle.
  • Citrate can cross the mitochondrial membrane. Remember you make citrate from condensing it with oxaloacetate and acetyl CoA.
  • There is another enzyme called a citrate-cleavage enzyme, which will spit out the acetyl CoA unit and oxaloacetate is generated.
  • This malate to pyruvate shuttle allows you to produce reducing equivalents that you will need to synthesize the fatty acids.
  • Just look at the malate to pyruvate shuttle and the citrate shuttle in order to get our substrate and our reducing equivalents from the mitochondria.
  • This makes sense why it was designed this way because if you have excess citrate in your mitochondria you have funneled down lots of acetyl CoA units. You must have eaten a big meal and a lot of glucose. These acetyl CoA units are coming from glucose. We don’t need them for energy, so we are going to ship them out into the cytoplasm and store them as fat.
  • The organ in which this takes place is primarily the liver (fatty acid biosynthesis machine of our body)

Controls for fatty acid biosynthesis

  • What controls fatty acid biosynthesis?
  • Remember we said that fatty acid biosynthesis is controlled at the first enzyme/step in the pathway, the acetyl CoA carboxylase, the enzyme that makes malonyl CoA from acetyl CoA.
  • All of the greens on the slide will be the things that stimulate this enzyme.
  • All of the reds on the slide will be things that stop or inhibit it.
  • The mechanism of activation or inactivation is by phosphorylation, in this case the kinase phosphorylates this carboxylase to produce an inactive carboxylase.
  • As we discussed before, for every kinase there must be a phosphatase to turn it back to the original state. These things will either stimulate the phosphatase or the kinase.

Activators & Inhibitors of Acetyl CoA Carboxlase

  • Things that keep the enzyme in its active form:
  • Citrate: store some substrate for fat
  • Insulin: will be high after a meal high in glucose; when insulin levels are up you want to activate your carboxylase and store your substrate as fat
  • If you have enough fat already, palmitate will shut down the carboxylase or stimulate the kinase to make an inactive carboxylase.
  • amp is a signal for low energy
  • If you are low on ATP do you want to make fat?
  • No! You want to use your ATP to do other things
  • Glucagon is a hormone that increases in starvation & you want to utilize fat.
  • Epinephrine/adrenaline – fight or flight mechanism
  • You don’t want to make fat in some situations.

Acetyl CoA carboxylase activity: phosphorylated vs. dephosphorylated

  • Citrate is interesting because even the inactive carboxylase, which is phosphorylated can be partially activated by citrate.
  • This is the highly phosphorylated enzyme without much citrate it has relatively low activity compared to the dephosphorylated/activated one.
  • As citrate concentrations increase, even the inactive/phosphorylated enzyme then increases in activity. The dephosphorylated one doesn’t do as much as far as percentage of increased activity, but for the phosphorylated one it does, it activates it.

New Notes

  • What happens during starvation?
  • We see that glucagon goes up, blood glucose goes down, but let’s focus on two of the things that we are interested in:
  • plasma free fatty acids go way up
  • blood ketone bodies also go up.
  • Plasma free fatty acids bound to serum albumin are mobilized from adipose stores. They are now supplying me with energy. I am mobilizing my fat from adipose stores in order to supply energy to other organs.
  • Why do we need to produce ketone bodies? A lot of organs can oxidize fat. In fact, most organs can. They can use fat for energy during starvation.
  • One notable exception is the most important organ that you have – brain
  • Your brain does not do fatty acid oxidation.
  • The brain has been designed to use ketone bodies, which have been formed from fat so the brain can also get some energy from fat stores.
  • The increase in ketone bodies in plasma is also the hallmark of the ketogenic diet—the diet where you eat no carbohydrates. You force your body to mobilize fat stores and that produces these ketone bodies
  • What are the ketone bodies?They are not actually bodies.
  • From fat you can get 2 acetyl CoA units that can be condensed into aceto-acetyl CoA…add another 2 carbon unit and get beta-hydroxymethylglutaryl CoA.
  • This is one of the substrates that begin cholesterol biosynthesis. We are going to break this substrate into our ketone bodies.
  • This is a really strange type of metabolism. You are taking two acetyl groups to make a 4-carbon unit. You are taking another one to make a 6-carbon unit, then you are cleaving off 2 to get you back to the 4-carbon compound.
  • What happens is you remove the CoA group. That is the strategy.
  • Now we have aceto-acetate, which is the substance found in plasma when you are starving or you have a low-calorie diet. It is in equilibrium with beta-hydroxybutarate.
  • It also can be non-enzymatically converted to acetone. That is why if you smell a fruity chemical on a patient’s breath, they are blowing off acetone. That appears on the breath of the patient. You will be able to tell who is on one of these ketogenic diets, because they will have this acetone coming off of their breath.
  • Aceto-acetate: a ketone compound is taken up by the brain and is metabolized to 2-acetyl CoA units and the brain can then put those CoA units into the TCA cycle and get energy from fat.
  • Essential fatty acids: Focusing on linoleic because that one can be metabolized to arachidonic.
  • Arachidonic acid is a C-20 fatty acid. This acid is the precursor of precursor of prostaglandin and is stored as phospholipids in membranes. It can be released from membranes.
  • Arachidonic acid is made by linoleic acid, which is a C-16 fatty acid. C-16 is converted to C-20. This is why linoleic acid is an essential fatty acid, because it is a precursor of arachidonic acid.
  • Arachidonic acid is a precursor of prostaglandins when it goes through the cycle oxygenase pathway or the prostaglandin synthestase pathway.
  • All these (on slide) are prostaglandins or thromboxanes.
  • When it goes through the lipoxygenase pathway, it creates leukotrienes.
  • Note the hormones-like compounds which exert various types of activities. Some lower fever. Some reduce pain. Some promote blood clotting.
  • Arachidonic acid didn’t show up well in this black box here.
  • You can either go to lipoxygenase pathway for leukotrienes or the cyclooxygenase pathway for these prostaglandins and thromboxanes.
  • The cycloxygenase reaction can be inhibited by aspirin, which is a covalent inhibition of the enzyme. Aspirin uses the acetyl-group to inhibit the enzyme. This is the reason that low dose aspirin therapy is used to prevent heart attacks. Many of the thromboxanes promote platelet aggregation. So if you block this pathway, you are much less likely to clot if you have a constricted coronary artery. The dose of aspirin about 80-90 mg opposed to the 300 mg doses that is marketed for pain and fever
  • Fish oils
  • Arachidonic acid is a C-20 acid (again)
  • Difference between this acid and acids that are found in fish oils
  • If we count from the omega-end, the last carbon of the arachidonic acid, our first double bond begins 6 carbons from the end of this acid. This is called an omega-6. Arachidonic acid is an omega-6 (6 refers to the position of the first double bond counting from the end).
  • Eicosapentaenoic acidis one of the main components of fish oils
  • Let’s look at where the first double bonds begins in this fish oil acid
  • This is called an omega-3 (count from the end)
  • These are extracted from cold water fish. All of them are omega-3 fatty acids. We looked at the enzymes which convert arachidonic acid to various prostaglandins and thromboxanes and leukotrianes. These enzymes act on omega-3 as well as omega-6. Eating a lot of omega-3 will displace omega-6 from the active site. These different prostaglandins and thromboxanes being produced instead of the normal ones.
  • What does that do? It turns out that one of the properties of these different prostaglandins and thromboxanes is that they are much less clot-promotting (platelet aggregating promoting). In fact, they are so much less, that the Eskimos don’t have a problem with heart attacks. They may have clogged arteries, but they will never throw a clot to block it. They will still get blood through it. The Eskimos who eat a lot of cold water fish (fish oil consumption) will die of cerebral vascular accidents (strokes) and they bruise easily. Sometimes clots are good, especially in the brain.
  • One researcher lived with the Eskimos and had their diet. He wanted to see how anti-clotting compound from these omega-3 were. How long would it take his blood to clot? The answer was one hour from when he pricked his finger.
  • If you have patients who eat fish oil what is going to happen to their membranes?
  • They will become what they eat. They will become more like fish membranes more than mammal membranes.
  • Note the two omega-3 acids in fish oils. At week zero, you have low concentrations of these, after 8 weeks of taking 3 grams of fish oil per day, what happens is the concentration of both of these concentrations goes up and the concentration of arachidonic acid goes down.
  • Same with phosphatidylethanolamine. Concentration goes up and arachidonic acid goes down. They are increasing their concentration in the membrane by displacing arachidonic acid in that membrane. Once they displace that acid, these can be mobilized and produce omega-3 analogs of all of the thromboxanes and prostaglandins.