The Citric Acid Cycle – Part II
Chapter 19 - pages 556-571Outline
• brief review of the citric acid cycle: so far… • the second half of the citric acid cycle
- regenerating oxaloacetate
• control and regulation of the citric acid cycle
• the role of the citric acid cycle in catabolism and anabolism
• after entering the matrix of the mitochondria, pyruvate is acted on by the PYRUVATE DEHYDROGENASE COMPLEX
- the PDC decarboxylates pyruvate and links what remains (acetyl) to coenzyme A - acetyl-CoA can then enter the citric acid cycle
• in step one of the citric acid cycle, the acetyl group from acetyl-CoA is transferred to oxaloacetate making citrate
• step two of the TCA cycle is the isomerization of citrate to ISOCITRATE
• step three of the citric acid cycle is the oxidative decarboxylation of isocitrate to α-ketoglutarate – CO2 is released
• step four of the citric acid cycle is another oxidative decarboxylation where α-ketoglutarate combines with CoA to make succinyl-CoA
• we reduced three NAD+ to NADH and released three CO2 molecules
The Citric Acid Cycle
• so, let’s pick up where we left off… at succinyl-CoA
• the bond between succinate and CoA (making succinyl-CoA) is broken through _____________ to yield succinate and CoA
- in the process, GDP is also phosphorylated to _______
• this reaction is catalyzed by succinyl-CoA _____________________ - this time named for the reverse reaction…
* side note: both synthases and synthetases MAKE NEW BONDS AND THEREFORE MAKE NEW MOLECULES
- the difference between them is that synthases do NOT require any energy from the hydrolysis
of a _________________________ bond (ATP) - synthetases do require this form of energy
• in this reaction the energy is coming from the hydrolysis of a phosphate that is part of the enzyme – SUCCINYL-CoA SYNTHETASE - itself
• of course, making a high energy molecule like GTP – phosphorylating GDP – takes energy and is
_________________________
- the energy to do this comes from the cleavage (hydrolysis) of the bond between succinate and CoA
- the ΔG is close to __________ and the universe is _______________________
• another enzyme, nucleosidediphosphate ______________ takes the new phosphate from GTP and uses it to phosphorylate ADP to ATP – “substrate level phosphorlyation”
• this is the only time in the citric acid cycle where we will make USEABLE energy - and it’s almost as though it’s an afterthought
- “Oh, look. Energy is being released…” • we’ve just made succinate
- as I told you on Wednesday, the rest of the citric acid CYCLE is dedicated to regenerating oxaloacetate
- coming back to the beginning so that we can cycle again
• there are three more steps to the citric acid cycle (steps 6, 7 and 8) which will convert four-carbon succinate into four-four-carbon oxaloacetate
• in step 6, succinate is oxidized to ______________________
- catalyzed by ______________________ dehydrogenase (you must know this enzyme – you’ll see why shortly)
• we mentioned on Wednesday that all of the reactions of the citric acid cycle occur in the mitochondrial matrix except for one…
- this is the one that doesn’t
• __________________ dehydrogenase is an ___________________ protein at the inner membrane of the mitochondria – what does this mean, again….?
• therefore, this reaction occurs at the inner membrane and not in the matrix itself
• also, as you can see, for the first time it’s NOT NAD+ that’s reduced in this redox reaction, but
FAD (reduced to FADH2)
- this FAD is actually covalently bound to the enzyme succinate dehydrogenase
- again, while FAD is being reduced, succinate is oxidized to _______________________ • succinate dehydrogenase is distinct in the citric acid cycle not only because it is the only
enzyme imbedded in the mitochondrial inner membrane - this enzyme also does double duty
- it is also part of the electron transport chain
• in step 7, water is added to fumerate (a ____________________ reaction) and this removes the double bond
- this yields MALATE
• this reaction is catalyzed by FUMARASE • malate is chiral
- only the ‘L’ form is made
• in step 8, we will regenerate ____________________________ and come back to the beginning – completing the cycle
• in addition to this being our last step of the citric acid cycle, it is also our last ___________ reaction of the citric acid cycle
• malate is oxidized to yield oxaloacetate while NAD+ is reduced to NADH + H+
• this reaction is catalyzed by MALATE ________________________________
The Citric Acid Cycle – in Summary
• when we look at the citric acid cycle overall – and include the conversion of pyruvate into acetyl-CoA – we see:
- the production of three molecules of __________
- one molecule of GDP phosphorylated to GTP (which is then used to make ________)
- _______ molecule of FAD is reduced to FADH2
- ___________ molecules of NAD+ are reduced to NADH + H+
- _________ from the cycle; one from pyruvate → acetyl-CoA
• although we only made one ATP from the cycle, and two from glycolysis, remember that this was all an investment
- we will reap large rewards in the next stages of aerobic respiration when we begin making ATP by the bucket-full
• with the citric acid cycle being so critical, how is it controlled?
Controlling and Regulating The Citric Acid Cycle
• the citric acid cycle, overall, is _______________________ with a ΔG of -44.3 kJ/mol • only one reaction is strongly endergonic, but it’s coupled to one of the most exergonic
reactions of the cycle…
- which step of the citric acid cycle comes after step 8…? - this coupling keeps the cycle always moving forward
• there are three control points in the citric acid cycle - that is, three enzymes of the cycle are regulated
• in addition to these three control points in the cycle, the _________ is also regulated adding a fourth control point
• why are we doing all of this…? - what are we trying to make…?
- when would it make sense NOT to do all this stuff…? • and, that’s exactly what regulates the citric acid cycle
- _________ inhibits the cycle for obvious reasons
- ___________ also inhibits the cycle… - why…?
• the PDC is activated by ADP – makes sense…
- if ADP levels are high, we’ve burned a lot of ATP and need more • in mammals, this regulation is achieved through phosphorylation
- when ATP levels are high/ADP levels are low, the PDC is phosphorylated by a ______________, slowing it down
- as ADP levels climb, and the need for the PDC returns, a ___________________ is activated, it dephosphorylates the PDC and allows it to once again be active
• remember, the kinase and phosphatase are part of the PDC itself
- but this is not necessarily feedback inhibition
• although we haven’t discussed it, acetyl-CoA can come from many different places • if you’re getting plenty of acetyl-CoA from fats and proteins, then there’s no need to use
pyruvate to make more – save it for later
• now, on to the three controls point within the actual cycle
• they are achieved through the regulation of citrate synthase, isocitrate dehydrogenase, and the α-ketogluterate dehydrogenase complex
• citrate synthase is inhibited by: ATP, _____________, ___________________, and its own product – citrate
- the first two inhibitors keep this step sensitive to the energy needs of the cell - the second two inhibitors provide information about the rate of the cycle itself
- is it backing up…?
- slow down and let everyone else catch up!
• isocitrate dehydrogenase is activated by ADP and ____________ - same logic applies; different mechanism
• if ADP levels are high, then the cell recently burned through a lot of ATP
• if ____________ levels are high, then the cell is NOT about to make more
• those two conditions mean the citric acid cycle must pick up the pace and feed electrons on to the electron transport chain so we can crank out the ATPs
• isocitrate dehydrogenase is an __________________________ enzyme • α-ketoglutarate dehydrogenase is inhibited by ATP and NADH
- it is also inhibited by ____________________
• you may remember that we discussed how the PDC and α-ketoglutarate dehydrogenase are very similar
- they are similar in their regulation as well
• α-ketoglutarate dehydrogenase activity is regulated by kinases and phosphatases working in opposition
• although it is no surprise at this point…
The Role of the Citric Acid Cycle in Catabolism • we eat EVERYTHING!
- polysaccharides, proteins, lipids, etc.
• first, these large polymers are broken down into smaller, simpler subunits
• many of these subunits (monomers) can be converted into ___________________ and enter the citric acid cycle
• but, LOOK!!!
- some amino acids are converted into citric acid cycle intermediates… - this can speed up metabolism
The Role of the Citric Acid Cycle in Anabolism
• I have also already mentioned to you that the citric acid cycle plays a large role in ‘making things’
things, then we run the risk of shutting the cycle down - certain death to the cell
• the citric acid cycle must be ________________________________ - we’ve just seen one way how that can be done…
- but how can we replenish the key intermediate…?
- _____________________________
• the enzyme pyruvate carboxylase can make _______________________ from pyruvate (we encountered this enzyme in gluconeogenesis)
• this allows the citric acid cycle to give up some intermediates to anabolism, while still being replenished and kept on running
• acetyl-CoA is the starting point for lipid anabolism
• much of this acetyl-CoA comes from the consumption of ________________
- remember, ____________ to pyruvate in glycolysis and pyruvate to acetyl-CoA by the PDC
- it all traces back to ______________
• animals cannot convert lipids to carbohydrates, but they (we…) can convert carbs to lipids - we didn’t need a biochemistry class to tell us that…
• intermediates of the citric acid cycle are also precursors for amino acid biosynthesis
• oxaloacetate can be made into ___________________ by a single reaction
- _________________ can then be used to make many different amino acids as well as nucleotides and other nitrogen-containing molecules
• α-ketoglutarate can be made into __________________ by a single reaction
- __________________ can be used to make most of the remaining amino acids that did not derive from aspartate
• succinyl-CoA is the primary precursor used for making ___________ • it’s crazy how important the citric acid cycle is!!!
A Brief Point: why is the Citric Acid Cycle part of aerobic respiration?
• oxygen was not required for any of the reactions of the citric acid cycle - yet it is the defining pathway of aerobic respiration
- why…?
• everything done in the citric acid cycle was done so that ATP can be made - and making this ATP requires oxygen (as we will soon see)
• therefore, with no oxygen, no ATP would be made and all that the citric acid has done would be in vain
• so, although the citric acid cycle itself does not require oxygen, the grand goal of the citric acid cycle – making ATP – does
- therefore, this is all part of aerobic respiration
Summary
• as ADP levels climb, and the need for the PDC returns, a ___________________ is activated, it dephosphorylates the PDC and activating it
• ________________ also inhibits the PDC
• citrate synthase is inhibited by: ATP, ________________, ____________________, and its own product – citrate
• isocitrate dehydrogenase is activated by ADP and ____________
• α-ketoglutarate dehydrogenase is inhibited by ATP and ____________
- it is also inhibited by ________________
• many subunits (monomers) can be converted into ___________________ and enter the citric acid cycle
- some amino acids are converted into cycle intermediates
• the enzyme pyruvate carboxylase can make _____________________ from pyruvate
• oxaloacetate can be made into _________________ by a single reaction
- __________________ can then be used to make many different amino acids
• α-ketoglutarate can be made into ____________________ by a single reaction
- _____________________ can be used to make many different amino acids • that’s it for exam #2…
