Citric Acid Cycle and Oxidative Phosphorylation Regulation of Metabolism Flashcards
cellular respiration overview and 3 major stages
cells consume O2 and produce CO2
complete oxidation of glucose (–> CO2 & H2O) provides more energy (ATP) than glycolysis (glucose –> pyruvate), also captures energy stored in lipids and AA
stage 1: acetyl CoA production
stage 2: acetyl CoA oxidation
stage 3: e- transfer and oxidative phosphorylation
3 stages of cellular respiration
1: oxidation of FA, glucose, and some AA to produce acetyl CoA, generates ADH + NADH, final product acetyl CoA, e- donor is NADH.
2: oxidation of acetyl groups in the citric acid cycle, includes 4 steps in which e- are abstracted. remaining CO2 are released from CHO, AA and FFAs
3: oxidative phosphorylation, generates vast majority of ATP during catabolism.
e- carried by NADH and FADH2 are funnelled into a chain of mitochondrial E0 carriers, ultimately reducing O2 to H2O, this e- flow drives the production of ATP
chemiosmotic mechanism for ATP synthesis
in eukaryotes, stages 2 and 3 are localised to the mitochondria, glycolysis occurs in the cytoplasm
- citric acid cycle occurs in the mitochondrial matrix
- oxidative phosphorylation occurs in the inner mitochondrial membrane
conversion of pyruvate to acetylCoA
final step in stage 1 of respiration, highly thermodynamically favourable, irreversible reaction.
- function of CoA is to accept and carry acetyl groups
- net reaction: oxidative decarboxylation of pyruvate, first carbons of glucose to be fully oxidised (released as CO2)
catalysed by the pyruvate dehydrogenase complex (PDC)
- requires 5 coenzymes
pyruvate dehydrogenase complex (PDC)
PDC catalysed confession of pyruvate to acetylCoA, PDC is a large multi enzyme complex.
advantages of multi enzyme complexes: short distance between catalytic sites allows channeling of substrates from one catalytic site to another, minimises side reactions, regulation of activity of one subunit effects the entire complex
conversion of pyruvate to acetylCoA
highly favourable, irreversible reaction, generates NADH, produces CO2
role of the citric acid cycle
At each turn of the cycle, three NADH, one FADH2, one GTP (or ATP), and two CO2 are released in oxidative decarboxylation reactions (reversible reactions)
events of the citric acid cycle
Step 1: C-C bond formation between acetate (2C) and oxaloacetate (4C) via condensation to make citrate (6C)
& CoA-SH
Step 2: Isomerization via dehydration/rehydration converts citrate to isocitrate
Steps 3 & 4: Oxidative decarboxylations to give 2 NADH and CO2, regulated by [ATP]
Step 5: Substrate-level phosphorylation to give GTP (GTP converted to ATP)
Step 6: oxidation of alkane to alkene, via dehydrogenation to give FADH2
Step 7: Hydration across a double bond, product concentration kept low to pull reaction forward
Step 8: oxidation of malate and regeneration of oxaloacetate via dehydrogenation to give NADH
net result of the citric acid cycle
acetylCoA + 3NAD+ + FAD + GDP + Pi + 2H2O –> 2CO2 + 3NADH + FADH2 + GTP + CoA + 3H+
Net oxidation of two carbons to CO2, equivalent to two carbons of acetyl-CoA
Energy captured by electron transfer to NADH and FADH2, Generates 1 GTP, which can be converted to ATP
Completion of cycle
regulation of the citric acid cycle
Regulated at highly thermodynamically favorable and irreversible steps
PDC (pyruvate to Acetyl CoA),
Citrate Synthase, IDH, and KDH (steps 1,3 & 4, CAC)
General regulatory mechanism
o activated by substrate availability
o inhibited by product accumulation
Overall products of the pathway are NADH and ATP
affect all regulated enzymes in the cycle
- activators: NAD+ and AMP
- inhibitors: NADH and ATP
energy from reduced fuels is used to synthesise ATP
Carbohydrates, lipids, and amino acids are the main reduced fuels for the cell.
Electrons from reduced fuels are transferred to reduced cofactors NADH or FADH2.
In oxidative phosphorylation, energy from NADH and FADH2 is used to make ATP.
NADH & FADH2 transfer e- gained while oxidising other molecules to the Electron Transport Chain (ETC)
key electron carriers
NAD+: nicotiadamide adenine dinucleotide
- phosphorylation: NADP+
- reduction to NADH and NADPH
- accepts H+ ion from an oxidisable substrate
FADH: flavin adenine dinucleotide
NADH and FADH2 are the main products of glycolysis and the citric acid cycle
oxidative phosphorylation utilised the reduced co-enzymes to produce ATP via ETC
NADH = 2.5 ATP FADH2 = 1.5 ATP
chemiosmotic theory
ADP + Pi –> ATP
highly thermodynamically unfavourable
- phosphorylation of ADP not a result of a direct reaction btwn ADP and high energy phosphate carrier
- energy need to phosphorylate ADP is provided by the flow of protons down the electrochemical gradient
- energy released by e- transport is used to transport protons against the electrochemical gradient
energy coupling requires a membrane that is impermeable to ions, is either the plasma membrane in bacteria or inner membrane in mitochondria
- membrane must contain protons that couple the downhill flow of e- in the e- transfer chain with the uphill flow of protons across the membrane
- membrane must contain a protein that couples the downhill flow of protons to the phosphorylation of ADP
NADH and FADH2 movement in membrane
Electrons move spontaneously through a chain off membrane-bound carriers, the respiratory chain, driven by the high reduction potential of oxygen and the relatively low reduction potentials of the various reduced substrates (fuels) that undergo oxidation in the mitochondrion. Electron flow creates an electrochemical potential by the transmembrane movement of protons and positive charge. This electrochemical potential drives ATP synthesis by a membrane-bound enzyme, ATP synthase, that is fundamentally similar in mitochondria and chloroplasts, and in bacteria and archaea as well
structure of mitochondrion
double membrane leads to four distinct compartments
- outer membrane: porous, allows passage of metabolites
- inter membrane space (IMS): higher [proton], low pH
- inner membrane: impermeable, protein gradient across it, location of ETC complexes, convolutions called cristae serve to inc the SA
- matrix: location of citric acid cycle and parts of lipid and AA metabolism, lower [proton] and higher pH
ETC
located in the inner mitochondrial membrane, long series of specialised acceptor and donor molecules
made up of a series of e- carriers, 3 of which act as proton pumps
as e- pass along the chain they fall to successively lower energy states - energy released pumps protons out of the matrix into the intra membrane space and creates a proton gradient
ADP + Pi –> ATP, O2 reduced and combines with H2 to produce H2O
summary of e- transport
Complex I –> Complex IV
1NADH + 11H+(N) + ½O2 –> NAD+ + 10H+(P) + H2O
Complex II –> Complex IV
FADH2 + 6H+(N) + ½O2 –> FAD + 6H+(P) + H2O
Difference in number of protons transported reflects differences in ATP synthesized.
proton motive force
proteins in the ETC created the electrochemical proton gradient via 3 processes
- actively transporting protons across the membrane (complex I and IV)
- chemically removing protons from the matrix (reduction of CO2 and O2)
- releasing protons into the inter membrane space (oxidation of H2)
the inner mitochondrial membrane separates two compartments of different [H+], resulting in differences in chemical concentration (∆pH) and charge distribution (∆ψc) across the membrane. The net effect is the proton-motive force (∆G)
inhibitors of the ETC disrupt oxidative phosphorylation
method determining the sequence of e- carriers, measures the effects of inhibitors of e- transfer on the oxidation state of each carrier.
in presence of e- donor and O2, each inhibitor causes a pattern of oxidised/reduced carriers
regulation of oxidative phosphorylation
primary regulation by substrate availability (NADH and ADP/Pi)
inhibitor of F1 component of ATP synthase: prevents hydrolysis of ATP during lower O2, only active at lower pH, encountered when e- transport is stalled
inhibition of ox-phos leads to NADH accumulation, causes feedback inhibition cascade up to PFK glcolysis
control of metabolic pathways
- availability of substrates
- allosteric activation and inhibition of enzymes
- covalent modification of enzymes
- induction & repression of enzyme synthesis
integration of metabolism
- metabolic processes must be coordinated
- opposing pathways are not operating simultaneously
- cells must respond to constant changes: external (nutrients) and internal (growth and reproduction)
- multi-cellular organism cells must cooperate
- simplified by division of labour between tissues
- different pathways operate in different tissues
tissues involved with metabolism
RBC
brain, skeletal and cardiac muscle, adipose tissue, liver and red blood cells
RBC simples cellular metabolic machinery, no nucleus.
glycolysis –> pyruvate –> lactate via the pentose phosphate pathway (PPP)
- no mitochondria so no citric acid cycle/oxidative phosphorylation
Major role transport of Hb/O2
No expenditure or gain of energy all metabolic activity is directed @ providing energy for the correct ion balance. Pumping out of Na+ in exchange for K+
End product of glycolysis is Lactate - Glucose used in the pentose phosphate pathway provides NADPH to keep glutathione in a reduced state - removes peroxides & H2O2 cause irreversible damage to cell membrane.
brain tissue/metabolism
20% of resting O2 consumption, most E used for maintaining membrane potential for nerve impulses
glucose is primary fuel, requires steady supply
- extended fasting gradually switches to ketone bodies
acute low BGL –> coma –> death
glycolysis, PPP, CAC
- No gluconeogenic enzymes (can’t make glucose)
- no No β-oxidation machinery
- Brain takes up glucose by mediated transport - insulin-independent mechanism
- Glycolysis in brain yields pyruvate which can be completely oxidised to CO2 & H2O
- Pentose Phosphate Pathway is active in these cells - generates NADPH keeps glutathione in reduced state.
muscle tissue/metabolism
major fuels: glucose, FA and ketone bodies, rested and well fed stores 1-2% of its mass as glycogen.
- glycogen more rapidly mobilised than fat
- glucose can be metabolised anaerobically (FA cannot)
- muscle cannot export glucose
- muscle carbohydrate metabolism serves only muscle
- extreme exertion: muscle contraction is anaerobic, driven by ATP hydrolysis
- at rest: sk muscle users 30% of body O2
- heavy load: 25x more O2 used, higher demand for ATP
muscle tissue/metabolism & lactate
maximum flux of glycolysis exceeds CAC, so much of the glucose is degraded anaerobically to lactate, muscle fatigue caused by pH drop caused by lactate build-up
This explains why lactate has to travel from the muscle to the liver. Excess lactate produced in muscle goes to Liver - Cori cycle
heart tissue/metabolism
Heart is a muscular organ
- Continuous action - different to skeletal muscle
- Relies entirely on aerobic metabolism
- Has many mitochondria (40% of cytosol)
- Metabolises - glucose, pyruvate, lactate, fatty acids, ketone bodies
- rest: fuel is FA
- heavy work: small glycogen store, converted to glucose
adipose cells/metabolism
function to store and release FAs, distributed in skin, abdomen and muscle
- FA storage from circulating lipoproteins –> triacylglyerols
- inc glucose –> inc insulin –> FA mobilisation
- insulin –> glucose uptake –> glycerol-3-phosphate –> triacylglycerol
- lipase –> FFA + glycerol during fasting
- Intakes glucose by insulin dependent mechanism
- Pyruvate is oxidised to Acetyl CoA which is used in the de novo synthesis of fatty acids
- Adipose tissue has the capacity for gluconeogenesis and glycogenolysis - much more limited than in muscle & heart - therefore fatty acid synthesis is favoured
liver - glucose
- Central metabolic clearing house
- Acts as a blood glucose buffer
- Takes up and releases glucose in response to BGL & hormones
- Converts to glucose-6-phosphate: glucose, glycogen, Acetyl CoA, Pentose Phosphate Pathway
- Contains enzymes for gluconeogenesis
liver - FA
- Central metabolic clearing house
- Can synthesise or degrade Fatty Acids
- During High Metabolic Demand (ie low fuel), Can synthesise ketone bodies but not use them (Lacks enzyme for reconversion) and FA are preferred fuel
- During Low Metabolic Demand (ie high fuel): Fatty acids –> TAG –> adipose tissue
liver - AAs
- Amino Acids are metabolic fuels
- Can be completely oxidized to O2 & H2O
- Converted to glucose
- Converted to ketone bodies
liver/metabolism
- liver has the greatest number of ways to utilise glucose
- takes up glucose by insulin (independent mechanism)
- uses PPP extensively for NADPH production: required for reductive synthesis, reduced glutathione, Er enzyme systems, provides ribose phosphate for nucleotide
- uses glucose for glycogen synthesis
- glucose used the glucoronic pathway (drug and bilirubin detoxification)
- glycolysis: pyruvate –> acetyl CoA –> CAC
- use Acetyl CoA for de novo fatty acid synthesis
- Liver can convert lactate, pyruvate, glycerol, alanine into glucose by gluconeogenesis to meet the needs of other cells in the body.
cori cycle
- Anaerobic glycolysis in Muscle
- Pyruvate –> Lactate - transported in blood
- Lactate –> Liver –> pyruvate –> glucose/glycogen
- Rescues Lactate & Recovery of glucose
- Object of anaerobic glycolysis not to produce lactate but to re-oxidise NADH & thus permit continued ATP production from glycolysis.
cori cycle increased lactate - impact on muscle
- Increased Lactate DH in muscle therefore NADH is rapidly re-oxidised, this leads to increased rate of glycolysis
- Useful in fight & flight situation - rapid exercise means that energy requirements outstrip O2 supply.
- Anaerobic metabolism - ie glycolysis has the advantage that although only 2ATP are generated per molecule of glucose (as opposed to 38ATP with complete oxidation in presence of O2) Relatively vast amounts of glucose can be broken down - difference between being eaten or not.
- Short time later, increased heart rate, dilation of blood vessels increase O2 supply and oxidative regeneration of NAD+ increases
- Excess lactate produced in the muscle must travel to the liver where it is converted to pyruvate and can then be used to generate glucose via gluconeogenesis
- Effects
1. rescues lactate for further use
2. counteracts lactic acidosis (too much lactate => dec blood pH)
metabolic cooperation between sk muscle and the liver - cori cycle
Extremely active muscles use glycogen as their energy source, generating lactate via glycolysis. During recovery, some of this lactate is transported to the liver and converted to glucose via gluconeogenesis. This glucose is released to the blood and returned to the muscles to replenish their glycogen stores. The overall pathway (glucose –> lactate –> glucose) constitutes the Cori cycle
fuel movement during starvation
- Initially liver breaks down glycogen –> glucose
- Then gluconeogenesis (GNG) is required. The liver needs substrates from other tissues to form glucose using GNG.
- ie glucogenic amino acids mostly from the muscle (muscle switches to using FFA’s and ketones for energy, spares use of glucose)
- After a few days of fasting most of the energy requirements of the the body are met by fat catabolism = glucose sparing mechanism. TAG’s in adipose tissues are broken down to FFAs and these circulate to the Liver and muscle.
- Remainder is supplied by glucogenic amino acids
- The brain adapts to using ketones
ketosis in starvation
- Liver makes inc Acteyl CoA when fat is mobilised & glucose is reduced
- inc Acteyl CoA normally –> TAG synthesis but if glucose is dec –> dec TAG
- Therefore excess Acteyl CoA
- -> ketones –> leads acidosis in starvation
- The liver maintains glucose output using the amino acids for gluconeogenesis that come from muscle
tissue communication
Occurs via
- Availability of substrates
- Hormones
- Nervous System
to the liver, adipose, muscle and brain
integration of metabolism
- Metabolism also under hormonal control
- Insulin –> uptake of glucose, dec glucose synthesis
- Glucagon –> release of glucose inc glucose synthesis
- Epinephrine –> inc glucose synthesis
