Glycolysis & gluconeogenesis: In-too-deep edition. . . (For those who like details!)

You’ve gotta spend (energy) money to make (energy) money! The first steps of breaking down blood sugar for energy (glycolysis) may seem counterintuitive if you just try to memorize them - which I hope you don’t, but rather focus on the logic! - (it starts with spending energy in an “investment phase”!), but learning about METABOLISM need not bring pessimism! 
 
blog form: bit.ly/metaboli...
if you want the graphics: bit.ly/glycoly... (and please cite the bumbling biochemist if you use them - thanks!) 
GLYCOLYSIS starts with an “investment phase” in which energy is spent followed by a “payout phase” in which energy is produced. In GLYCOLYSIS (a catabolic process) you split the sugar glucose (which has 6 carbons (6C)) into 2 molecules of pyruvate, each of which have 3 carbons (3C). This nets you 2 ATP (energy “arcade tokens”) and 2 NADH (energy “IOUs”)(more below), and those 2 pyruvate, which can be further broken down in the citric acid cycle (aka TCA or Krebs cycle). 
 
ENERGY INVESTMENT PHASE 
 
STEP 1: enzyme: hexokinase; reaction: glucose + ATP → glucose-6-phosphate (G6P) 
a kinase is a phosphate adder, and in this step phosphate is added to the “6th” of glucose’s 6 carbons 
this is the first of the “irreversible” steps - you start by putting down an energy-money downpayment 
it also helps with the committing because the charge makes it “impossible” for glucose to get out of the cell - what with the cell’s fatty membrane and all - so the glucose gets trapped 
 
STEP 2: enzyme: phosphoglucose isomerase; reaction: glucose-6-phosphate ⇆ fructose-6-phosphate 
isomers are different arrangements of the same atoms so, as the “isomerase” name suggests, what goes on in this step is just some rearrangement - specifically carbonyl (C=O) that glucose ends with gets shifted over, so that there’s an OH on the end. This might seem weird but just wait - it’s gonna be important for… 
STEP 3: enzyme: phosphofructokinase; reaction: fructose-6-phosphate + ATP → Fructose-1,6-bisphosphate 
this is the second “irreversible step” - and it involves another phosphate adding - this time to the 1st carbon which you graciously made available in step 3 
why the second “downpayment”? if you look at the structures you’ll see that you now have a phosphate on both ends, so when you split it in half each half gets one 
the kinase involved is highly regulated to help regulate glycolysis as a whole 
STEP 4: enzyme: fructose bisphosphate aldolase; reaction: fructose-1,6-bisphosphate ⇆ dihydroxyacetone phosphate (DHAP) + glyceraldehyde-3-phosphate (G-3-P) 
this is the splitting I was talking about - you take a 6C thing and split it into two 3C things 
the “halves” aren’t identical in the beginning (they have carbonyls in different spots) but, that’ll get sorted out with the help of another isomerase in… 
 
STEP 5: enzyme: triose phosphate isomerase; reaction: DHAP ⇆ G-3-P 
those unidentical halves have the same atoms just arranged a little different - they’re isomers - so, with the help of an isomerase, you can shift between them 
only G-3-P can be directly used in the next step so, even though the reaction’s easily reversible, since you keep taking away the G-3-P, if you want to convert something, you’ve gotta convert the DHAP since there’s “no” G-3-P available. so eventually all the DHAP gets turned into G-3-P so you have 2 identical G-3-P’s entering the…. 
 
PAYOFF PHASE 
 
This is where you finally start making energy money (ATP) & energy IOUs (NADH). And the important thing to remember is that you’re going in with 2 copies of the G-3-P so each of the reactions gets “multiplied by 2” 
 
STEP 6: enzyme: glyceraldehyde-3-phosphate dehydrogenase; reaction: glyceraldehyde-3-phosphate + NAD+ + Pi ⇆ 1,3-bisphosphoglycerate + NADH + H+ 
this is a kinda “weird” step because it adds a phosphate but a free-floating one (so it doesn’t cost any ATP) - the phosphate gets added to the “other end” so you now have one on each end again 
the energy for the adding comes from the coupled redox reaction of NADH reduction & glyceraldehyde-3-phosphate oxidation, which is exergonic (energy-releasing) because the NAD+ wants the electrons more 
this is your first IOU payment - it reduces NAD+ to NADH which can then be “cashed in” for ATP later in oxidative phosphorylation 
 
STEP 7: enzyme: phosphoglycerate kinase; reaction: 1,3-bisphosphoglycerate + ADP ⇆ 3-phosphoglycerate + ATP 
our first real payout! 
don’t get confused by the enzyme name - before we had kinases using ATP to add phosphates, but here we’re adding a phosphate to ADP - and we’re getting that phosphate by taking off the one we just added in step 6 (the name refers to the reverse reaction) 
 
Finished in comments

[@thebumblingbiochemist]

STEP 8: enzyme: phosphoglycerate mutase; reaction: 3-phosphoglycerate ⇆ 2-phosphoglycerate 
- this is another isomerase, but I guess they thought “mutase” sounded cooler or something - but “all” you’re doing here is shifting the phosphate to the middle C 

STEP 9: enzyme: enolase; reaction: 2-phosphoglycerate ⇆ phosphoenolpyruvate (PEP) + H₂O 
- making PEP is a kind of “prep” - when you kick out that water you make a double bond between 2 of the carbons - making it a really awkward situation for that middle carbon - this molecule is really unstable 
- so the phosphate is now attached to a carbon that doesn’t really want it and can better survive without it - so that phosphate can more easily get removed in… 
 
STEP 10: enzyme: pyruvate kinase; reaction: PEP + pyruvate kinase + ADP → pyruvate 
- the last step & final payout of glycolysis 
- involves another kinase named for the reverse reaction 
- because PEP is so unstable, it’ll easily give up that phosphate to an ADP that pyruvate kinase helps it meet 
- and because of how unstable PEP is, although you’re not spending energy money, the reaction’s really unlikely to go backwards. 

So you do all that for each copy of G-3-P you got from the investment phase. So, you end up with 2 X NADH (from step 6), 2 X ATP from step 7 and 2 x ATP from step 8. And you used 2 ATP in the investment phase. So, on net you have: 2 NADH + [(-2) + 2 + 2 = 4] ATP. And you also have those 2 pyruvates which can get further processed for more energy. 
 
Gluconeogenesis is the “reverse” of glycolysis but it’s not a direct reverse because it has to “reroute” around the irreversible steps using different enzymes. It reverses step 10 (the de-pep-ing) in 2 or 3 steps - it’s a bit “steppy” because that pyruvate that got made in glycolysis gets shipped into the mitochondria. In the mitochondria, carboxylase converts pyruvate to oxaloacetate (at the cost of 1 ATP) & then phosphoenol-pyruvate carboxykinase converts that oxaloacetate. That can’t go through the mitochondrial membranes, so it first gets made into malate by malate dehydrogenase, then that malate goes into the cytoplasm where another malate dehydrogenase turns it back to oxaloacetate which can then be turned into to phosphoenolpyruvate (PEP) (at the cost of 1 GTP) by PEP carboxykinase in the cytoplasm. Alternatively, the PEP can be made in the mitochondria in some animals and transported out like that. 
 
It uses fructose 1,6-bisphosphatase to reverse step 3 (remove the second phosphate that was added to go from fructose 1,6-bisphosphate to fructose 6-phosphate. 
 
And to reverse the 1st step of glycolysis (initial phosphate-adding), glucose-6-phosphatase removes the phosphate to form glucose. This happens in the lumen of the endoplasmic reticulum (another membrane-bound compartment that’s often used for protein-modifying) and shipped out into the cytoplasm by glucose transporters. 
 
I learned some cool things about glycolysis in the book “For the Love of Enzymes” by Arthur Kornberg that president-elect of the IUBMB, Dr. Alexandra Newton, gave me. For example, the discovery of glycolysis came with the birth of “biochemistry” - and it came by accident - in a story not so well-known as the Pasteur and the Petri dish one about the discovery of the antibiotic penicillin being made by mold on a contaminated bacteria culture plate. 
 
Basically, a couple of guys in the late 1900s (1897 to be exact) - Hans & Eduard Buchner - thought there could be medicinal value to the stuff inside of yeast cells (cell-free yeast extracts). So they ground up & squished a bunch of yeast cells, filtered it and, to preserve it, added some sugar (hey - it works for jams right?!) Well, they found that the yeast extract started getting all bubbly - it was fermenting (fermentation is a process you can learn more about in yesterday’s post whereby NAD+ can be regenerated from NADH without going through oxidative phosphorylation) 
 
This was really surprising because, before this, it was thought that complex processes like this could only take place INSIDE OF LIVING CELLS - but this was just the cell extract! Further excitement came when studies with muscle extracts showed that lactic acid fermentation used a lot of the same reactions as the alcoholic fermentation that the yeast were doing. In fact, glycolysis is a nearly universal process in all sorts of organisms. Lots of pioneering biochemists joined in on the fun and, by 1940, the glycolytic pathway was figured out! Nowadays, we study reactions with purified proteins outside of cells all the time because it gives us better control
more on all sorts of metabolic stuff: bit.ly/bbmetabolism & ruclips.net/p/PLUWsCDtjESrHXBgulruKEOrNXQ21_0gyc
     
more about all sorts of things: #365DaysOfScience All (with topics listed) 👉 bit.ly/2OllAB0 or search blog: thebumblingbiochemist.com

[@tuppercareyd]

Thanks so much for these excellent videos!

[@thebumblingbiochemist]

Thank you! Glad you find them helpful!