16. The TCA Cycle Flashcards
What breakdown products can be fed into the TCA cycle?
What does this cycle need to run?
Breifly describe the TCA cycle.
Glucose (pyruvate), AAs (pyruvate), fats (beta oxidation produces acetyl CoA)
Constant supply of substrates and O2. Relies on ETC to regenerate its cofactors.
Acetyl CoA (2C) conbines with oxaloacetate (4C) producing citrate (6C). Lose 2 CO2 and regererate oxaloacetate. Produce reduced NAD and FAD, and 1 GTP along the way.
What is PDC (pyruvate dehydrogenase complex)?
Describe the journey of pyruvate through PDC.
3 stage enzyme. Essentially unidirectional under physiological conditions due to loss of CO2 = imp step of control. Uses cofactors e.g. TPP (thiamine pyrophosphate from vit B1).
1) Pyruvate dehydrogenase takes pyruvate, releases CO2 and attaches the 2C acyyl group to thimin pyrophasphate (TPP)(abosorbed from vitamin B1) to produce acyl-TPP
2) Transfer of 2C acyl group onto lipid amide arm of 2nd enzyme, then interacts with 2nd enzymatic site (CoASH) that takes CoA and attaches acetyl group to it, producing acetyl CoA
3) This reduced the lipid amide arm - 3rd enzyme oxidises lipoamide (and produces reduced NAD) so it can be reused
So this produces CO2, acetyl CoA, and reduced NAD
What 2 things control the PDC?
How does glucagon affect PDC?
1) Feedback inhibition: high levels of acetyl CoA and NADH inhibit PD
2) Covalent modification via phosphorylation by PDH kinase and PKA (phosphorylated PD is inactive)
Raises cAMP levels -> turns on PKA -> phosphorylation of PDC and turning off glycolysis because low sugar levels. Move to alturnative fuels and liver does gluconeogenesis instead. (Insulin dephosphorylates it = active via PDH phosphase)
Breifly describe the TCA cycle, including what is used up, and what is produced.
Is the TCA bidirectional? Why?
How many oxidation reactions occur in the TCA cycle?
How is GTP produced?
Acetyl CoA combines with oxaloacetate and goes around in a cycle to regenerate oxalacetate. 2C are lost as 2 CO2. Produce 3 reduced NAD (NADH) and 1 reduced FAD (FADH2), and 1 GTP (=ATP).
It is apart from 2 oxidation steps involving dehydrogenase enzymes (the steps where CO2 is lost). These steps are unidirectional so the cycle can only go one way even though a number of others could potentially go another.
4
Substrate level phosphorylation.
Describe how the reduced coenzymes generated in TCA, glycolysis and FA oxidation used?
What are UQ and Cytochrome C?
Describe what the ETC made up of, and how it works.
They feed e- into the ETC, down the electrode potential gradient to pump H+ from mitochondrial matrix to the inner membrane space. ATP is produced using the energy stored in this H+ gradient (PMF).
Small e- carriers that move e- between larger complex proteins in ETC.
4 complexes. NADH binds to complex I, deposits e<strong>-</strong> and gets oxidised, e- carried by UQ to complex III -> Cytc carries e- to complex IV -> e- flow through it and onto O2, producing H2O). FADH2 binds to complex II, deposits e- and gets oxidised, e- carried by UQ to compelx III -> Cytc carries e- to complex IV etc.
Complexes I, III and IV pump H+ across the membrane to generate a H+ gradient which is used to pump ATP. Complex II doesn’t pump H+ so get less H+ and ATP from FADH2 oxidation.
NB: NADH -> NAD+ + 2e-
What role do metal ions play in the ETC?
Why is energy produced from complexes I, III and IV in the ETC?
What is uncoupling?
What are the 2 ways of uncoupling?
Redox metabolism is dependant on minerals and trace elements e.g. Fe (complex I and II) and Cu (complex IV)
They allow change in redox potential (-ve e- move to +ve potential, and the more they move the more energy you can get out) - the change can be coupled and used to transport H+ across the membrane. (Complex II = no significant change in elextrode potential so unable to pump H+).
H+ gradient is coupled to ATP production in ETC, or ATP synthesis coupled to e- transport. Uncouple = provide alturnative route to dissipate the H+ gradient and doesn’t produce ATP.
1) Alturnative route for H+ to flow back through membrane instead of ATPsynthase
2) Uncouple charge gradient by letting ions flow through membrane to balance out the charge
How does non-shivering thermogenesis work?
Where is this particularly important in?
How is uncoupling used in natural antibiotics?
Uncoupling protein 1 (thermogenin) provides alturnative route for H+ to flow back across membrane and releases heat (energy makes heat instead of ATP)
Mitochondria of brown adipose tissue in newborns and small organisms (b/c high SA:V)
In bacteria ETC done over their own membrane, so if disrupt it = degeneration. Gramicidin forms 2 half channels in the bacterial membrane. Nigericin enables H+ to permeate membrane. Valinomycin enables dissipation of charge through movement of K+
How does DNP (dinitrophenol) effect the ETC?
Why was it considered for use in diet pills, and what is the problem?
Is membrane soluble so goes into innermembrane space of mitochondria, picks up H+, flows through membrane and deposits it on other side, and returns etc. = forms cycle that dissipates the gradient. Heat is produced b/c there’s energy in the gradient.
Glucose and fats can enter the TCA -> ETC to produce ATP, and if its used to produce heat, then we burn burn fat and glucose without producing any energy, so burn away calories as starved body starts using fats for energy. Problem: lethal dose v. close to theraputic dose (and energy made = heat). Used as pesticide/insecticide today. Treat overdose = cool pt down and manage symptoms!
What does overall TCA rate depend on?
What are dehydrogenases stimulated by in muscle?
Why is there not much control of the TCA?
What happens if there is a low availabilty of C?
NAD+ availability (which depends on ETC rate - thus linked to ATP/ADP ratio).
Ca2+ (b/c muscle contraction releases Ca)
Intermediate compounds used in other part of metabolism - to make AAs, nucleotides, as C skeletons for anabolism.
Can be topped up via pyruvate carboxylase.
Apart from the TCA cycle, what else can citrate be used for (when in excess)?
Will removing citrate from the cycle result in a lack of oxaloacetate?
FA can be broken down by beta-oxidation to form acetyl CoA - can this be used to make glucose?
How can glucogenic AAs feed into the TCA cycle beyond alpha-ketoglutarate?
Exported to cytosol and converted back to acetyl CoA and oxaloacetate. ACoA can be used for fat synthesis (FAs and cholesterol).
No - can top up cycle and produce more oxaloacetate: pyruvate carboxylase can produce oxaloacetate from pyruvate (as well as producing ACoA from pyruvate).
No - it can only be used for energy generation b/c lose 2C as go around TCA cycle which is the 2C in ACoA - no way of making oxaloacetate from it.
Alpha ketoglutarate goes on to produce oxaloacetate which can be used to synthesise glucose in gluconeogenesis.
How does hypoxia affect the ETC?
What 3 adaptation occur during (long-term) hypoxic events?
What are all metabolic adaptations controlled by?
How does it work?
O2 needed for complex IV. If no O2: e- back up. e- are reactive and contain energy. They can leak out of the complex (particuarly complex I which is close to surface) and interact with other chemicals in the cytosol to produce ROS which have excess e- and cause cellular damage (longterm).
1) Limit ATP use by switiching off non-essential cell functions
2) Improve anaerobic ATP production efficiency
3) Limit oxidative stress, providing protection against ischaemia i.e. produce enzymes and compounds that mop up ROS
HIF1 (hypoxia-inducible factor 1) - a transcription factor that results in altered gene expression e.g glycolytic genes upregulated in anaerobic conditions.
Normoxia: HIF1a subunit degraded. Hypoxia: HIF1a stabilised and binds to upstream elements of promotors (HREs)
How does HIF1 work?
What mitochondrial effect does it have?
Normal: HIF1 alpha picks up 2 OH groups -> provide protein binding site that targets HIF1a for degradation. Hypoxia: OH not added. Beta subunit binds to alpha -> interact with DNA and turn on genes for glycolysis, glucose transport (increase number of GLUT1) etc. as well as genes that increase Hb amount, and vascularisation to increase O2 to tissue.
Downregulates mitochondria respiration during hypoxia = promotes loss of mitochondria (autophagy) and supresses biogenesis (via PGC-1). Adaptive mechanism to prevent oxidative stress = less ROS produced.
