Bioenergetics, Glycolysis, Krebs cycle+ Electron Transport chain- Biochemistry Semester 2
What are Catabolic Pathways
They will release energy and involve the breakdown of molecules
What are anabolic pathways
These will require energy to take place and are involced in the synthesis of complex molcules from smaller molecules
Why is Glucose oxidation needed
It is needed for the production of free energy in the form of ATP and is also needed for the production of intermediates from glycolysis and the TCA cycle
What is oxidatove phosphorylation
Oxygen is used to make energy from carbohydrates and this is doen by aerobic respiration
How are Metabolic pathways characterised
By using the change in Gibbs free energy to characterise the steps, this is due to:
Life require energy so should be the main focus
Before all structural biologocal techniques it can be used to make reliable data
Individual Values are addtive for global overview and it can be applied for all pathways
What is delta G o'
This is the change in Gibbs free energy at pH 7 under standard conditions It can provide infomations about what happen during reacyion and is the amount of free energy between products and reactants
What does the Delta G value mean
If it negative then there is an exergonic reaction so energy is absorbed and if it is positive then there is an endogonic reaction so energy is released
How does Kc equilibrium constant effect metabolic pathways
If reaction lie to the lef then small Kc so is unreactive, if it lie in the middle then Kc is intermediate value and if lie to the right then Kc is large then it is reactove and a - Delta G so is spontaeneous
How is Delta G = 0 at equilibrium useful
Due to Delta G = -RT ln(Q)
Q= [Product]/[Reactant]
What is Dekta G dependent on
A constant term whos value depend only on the reaction taking place and alsoa a variable term that depend on the concentration of the reactant and products
Delta G is indicative of life being in a steady state rather than at equilibrium as in all living systems there is a negative delta G
What is le Chatelier Principle
Any Deviation from the equilibirum will cause a process that will restore equilibrium and many metabolic pathways are close to their equilibrium concentration but in the cell lie far away so the reaction is only 1 direction
How is delta G used in multiple step reaction
The Delta G values fro each of the steps are added togetehr and any of them can be + but if the overall reaction is Delta G - then the reaction is spontaneous and will proceed
How are Delta G and Equilibrium linked together
If Delta G is negative then lie in favour of products and so if + then lie in favour of reactants
If Delta G is large and - then reaction is irreversible and is large and + then reaction wont happen
What is the structure of glucose
The formula is C6H12O6 and it is present 36% of the time as alpha glucose, <1% as linear D glucose and 64% as Beta glucose
There is an aldehyde group on Carbon 1
All the hydroxyl groups are on the right hand side of the carbon chain except the hydrogen on Carbon 3
This is the structure when as D glucose
What are the 3 irreversible steps of glycolysis
This is due to large - Delta G making it irreversible
The first one is D glucose --> Glucose 6 Phosphate
The next one Fructose 6 Phosphate to Fructose 1,6 Bisphosphate
The final one is Phosphoenolpyruvate to Pyruvate
How is D glucose converted to Glucose 6 Phosphate
Need the enzyme hexokinase + Mg2+ and then aswell asATP hydrolysis to provide energy for this too happen
How is fructose 6 phosphate converted to fructose 1,6 bisphoshate
Need thr enzyme phosphofructokinase and ATP needs to be hydrolysed to provide energy for this to happen
The bisphosphate means that the 2 phisphate are added in a non sequential manner in different places
How is phosphoenolpyruvate convereted to pyruvate
This need to the enzyme pyruvate Kinase and ATP needs to be hydrolysed to provide energy for the reaction to take place
What is the structure of pyruvate
It is a 3 Carbon compound, on C1 there is a ester group on C2 there is a double bond O and then a methyl group
What are the roles of co-enzymes
These are non protein molecules in the enzyme and are used for the transfer of atoms and small molecules form one place to another
If the co-factor is organic then it will be called a coenzyme and a large number of them are from vitamins and metal ions
How are co-enzymes used in metabolism
They are used in the Catabolism of glucose so are thought to eb a co-substrate
They are involved in REDOX reactions when using NADH/NAD+
ATP is a coenzyme due to deliver a phosphate group
Why is NAD+ important
It can be reduced to NADH that is then needed in the ETC to make the proton gradient
Also needed for the reactiojn of Glyceraldhyde 3 phosphate to 1,3 diphosphoglycerate
How is Glyceraldehyde 3 phosphate converted to 1,3 diphosphoglycerate
There is the enzyme Phospoglycerate Kinase as well as ATP being hydrolysed to provide energy
NAD+ is then also reduced to NADH
What is ATP
It will tranfer energy between reactions but it wont store the energy due to very unstable
It is produced on demand by ADP phosphorylation
All phosphorylation reactions are endergonic and irreversible due to the adding of a phosphate group to a compound
What can ATO do to the equlibrium
It will couple exergonic and endergonic reaction to shift the equlibrium to favour the products in an endergonic reaction and to shift the products in an exergonic reaction
How can pyruvate be turned back to Phosphoemolpyruvate
It will first be carboxylate by pyruvate carboxylase + ATP hydrolysis to form oxaloacetate
Then the oxaloacetate will be decarboxylated to form the phosphoenolpyruvate p PEP- by the enzyme PEP carboxykinase
What is glucose metabolism
Gluvose is very adaptable and can meet the material and energy requirements of the cell unlike fatss
It has aan oxidation of Delta G -2838 = 95ATP but dont get this retunr due to use ATP in the process to oxidsize it
During glucose catabolism it is broken down in small steps and energy released in small amounts as chemical energy with ATP
What is glycolysis
This is where glucose is broken down to 2x Pyruvate + 2ATP + 2NADH + 2H+
Why are co-enzymes needed for glyolysis
They are needed for the addition of removal of phosphate groups that are delivered or to used to for ATP/ADP production
Also oxidation and reduction reactions use co-enzymes NADH/NAD+ that will add or remove protons and take them to ETC to create proton gradient
What are the 10 steps of glycolysis
Glucose os converted to Glucose 6 phosphate
Glucose 6 phosphate goes to Fructose 6 phosphate
Fructose 6 phosphate goes to fructose 1,6 phosphate
Fructose 6 phosphate is then broken down tb Dihyroxyiacetone phosphate + Glyceraldhyde 3 phosphate
Glyceraldehyde 3 phosphate is then converted to 1,3 bisphospoglycerate
1,3 bisphosphoglycertae is then converted to 3-phosphoglycerate
3-phosphoglycerate is then converted to 2 phosphoglycerate
This 2 phosphoglycerate then goes to Phosphoenolpyruvate- PEP
PEP is then converted to Pyruvate
What is the structure of glycerol
There is a 3 Carbon backobone that has fatty acids attached to each C
It is able to be metabolised to dihydracetone phosphaye that is then converted to Pyruvic acid
May also be used in gluconeogenesis to make glucose 6 phosphate
How is glucose converted to Glucose 6 phosphate
This use the enzyme Hexokinase + Mg2+ as well as ATP hydrolysis to provide energy
This is one of the 3 irreversible steps
The glucose can cross the cell membrane by specific transport proteins by diffusion
Once the phosphate added it cant bind to transport proteins so cant leave the cell
How is glucose 6 phosphate converted to Fructose 6 phosphate
This is by the enzyme phosphoglucose isomerase and it is reversible
How is fructose 6 converted to fructose 1,6 bisphosphate
There is the phosphorylation of the Carbon 1 and this is done by the enzyme Phospofructokinase
This is done so that when then fructose splits in half both 3 carbon molecules have a phosphate group
This is an irreversible reaction
How is fructose 1,6 bisphosphate converted to glyceraldehyde 3 phosphate - GAP + 1 x Dihydroacetone phosphate - DHAP
There is the enzyme aldolase that will split the 6 carbon fructose bisphosphate to 2x 3 C compunds that are different to each other
It is a reversible reaction
How is DAHP converted to GAP
All the DHAP will be converted to GAP as all the further steps go through GAP not DHAP
This is done by the enyzme troise phosphate isomerase and it is reversible
How is GAP converted to bisphosphoglycerate
This is doen by the enzyme GAP dehydrogenase and is reversible
It is driven by an oxidation reaction due to NAD+ will be reduced to NADH to then got the ETC for the creation of the proton gradient
How is 1,3 bisphosphoglycerate converted to 3 phosphoglycerate
There is the enzyme phospoglycerate Kinase and this is a reverisble reaction
There is also ADP goes to ATP due to the loss of a phosphate
How is 3phosphoglycerate converted to 2 phospoglycerate
This is done by the enzyme phosphoglycerate mutase and this a reversible reaction
How is 2 phosphoglycerate converted to phospheonolpyruvate
This is done by the enzyme enolase and is a reversible reaction with a water molecule coming off meaning it is a hydrolysis reaction
How is Phosphoenol pyruvate convereted to Pyruvate
This is done by the enzyme pyruvate Kinase and is a irreversible reaction and there is also ATP hydrolysis for the reaction to take place
What are the 3 irrervesible steps for glycolysis
Steps 1,3,10 are all irreverisble due to having a large - delta G value
What is produced during glycolysis
Produce 4 atp but 2 are used up so a net gain of 2ATP, also will produce 2 NADH and there are 4 phospjate used up
What are coenzymes
They are needed for an enzyme to become activated so it can carry out its catalytic properties once formed a holoenzyme which is the enzyme + Coenzyme
When the coenzyme binds to the enzyme it will change shape so is considerded the 2nd substrate
They need to be regenerated in order keep being part of the reaction
What is the structure of NAD+
There are many hydrogen bonds between the apartic acid and the ribose sugar ring system
There are + histidine and arginine side chains that interact with - phosphate group
Leucine backbone intercat with ribose sugar and phosphate group
There are lots of interactions go gold nicotinamide to prevent coenzyme moving around when trying to form holoenzyme
How does the enzyme glyceraldehyde 3 phosphate dehydrogenase work
It is the enzyme to turn glyceraldehyde 3 phosphate to 1,3 bisphosphoglycerate in glycolysis
There is the formation of thioether bond between substrate and enzyme
Next there is oxiadtion of thioether bond to a thioester bond by NAD+ that is reduced to NADH
The Thioester bond is then cleaved to supply the energy to the add the phosphate to form 1,3 bisphosphoglycerate
What happens to Pyruvate in aerobic conditions
Glycolysis will require the continual supply of NAD+ that can be met by the oxidation of NADH+ in the electron transport chain in the inner mitochondrial membrane
Pyruvate is able to enter the TCA cycle via the link reaction to then generate more NADH and ATP
What happens to pyruavte under anaerobic conditions
There is a limited supply of oxygen due to the organsim living in a confined space or situation with limited oxygenm#
There still needs to be NAD production from so then the pyruvate is converted to Lactate
Lactate can lead to lactic acid build up and oxygen debt in muscles
How is Pyruvate converted to Lactate
Pyruvate is converted to lactate and this will produce 2NAD+ and will use the enzyme Lactate Dehydrogenase
How does anaerobioc respirstion lead to alcohol production
Pyruvate is converted to 2 acetaldehyde by the enzyme pyruvate decarboxylase
The 2x acetaldehyde is then turned to 2x ethanol by the enzyme alcohol dehydrogenase
In the final step there are 2 NADH oxidised back to sNAD+
Why is the fermentation Pyruvate important
To produce many valuable good such as alcohol, dairy products
This would not be able to happen without glycolysis that could not happen without the regeneration of NAD+ from the ETC
All forms of fermenatation will oxidise NADH to regenerate NAD+ so that glycolysis can continue
Where does respiration take place
In Prokaryotic cells glycolysis, TCa and Fermentation all happen in the cytoplasmand the electron transport chain is in the cell membrane
In eukaryotic cells glycolysis and fermenatation take place in the cytoplasm, the TCA is in the mitochondria and the ETC is in the mitochondria membrane
What is the Fate of pyruvate in eukaryotic cells
The Pyruvate will leave the cytoplasm and enter the mitochondria to then enbetr the TCA cycle on the matrix
It will enter the mitochondria via porins to get into the intermembrane space but then hard to get through the inner membrane
There is a highly specific mitochondrial pyruvate carrier to get it through the inner membrane
NADH can pass through porins to enter the inner membrane
How is Pyruvate converted to Acetyl CoA
Done by the enzyme pyruvate dehydrogenase complex, and NAD+ is reduced to NADH
It is needed to be converted as acetyl is the substarte for the start of the TCA cycle
This is the link reaction
What is Coenzyme A
It is an acetyl carrier to active sites and will activate acetly, there is a thiol group that intercat with carboxyl acids and so is an acyl group carrier
There is an ADP molecule
uUsed for the synthesis of fatty acids when there is excess glucose
How is Pyruvate oxidised- Link Reaction
A carboxly group is removed from the pyruvate so then release CO2
NAD+ is reduced to NADH
Ann acetyl group is transferred to coenzyme A to make Acetyl CoA that is a 2 carbon compound to be fed into the TCA cycle
There are 2 pyruvate so lead to production of 2 NADH, 2 CO2 and 2 acetly CoA
What is the purpose of the TCA cycle
It is mainly for the productionof Co-factors and 1GTP (ATP) Molecule
Where is the TCA cycle in eukaryotic cells
Due to eukaryotic cells being large there is compartementalisation and so glycolysis and TCA cycle are in different location
In the TCA cycle one of the needed enzymes is an intergral membrane so then specific membrane has to be selected in the cell to produce the most energy efficiently
There is a close tie between the TCA to make the co-factors and the ETC where they are then oxidisded to release H+ and make the proton gradient
What is the evidence that mitochondria derived from bacteria
Size and shape are similar, They both replicate via fission and boht have circular DNA
Mitochondria make their own ribosomes to make their proteins and the ribosomes are more similar to bacteria than eukaryotic cells
Both have inner and outer membrane with porins
What does the crisate do in the mitochondria
They are invaginations of the membrane into the matrix area in order to increase surface area, in the membrane there are large protein levels and the ETC that produce the most energy so then a higher surface area lead to greater energy output
What is the Link reaction
The conversion of Pyruvate to Acetyl CoA by the enzyme Pyruvate dehydrogenase complex
There will be the Reduction of NAD+ to make NADH, the decarboxylation of the pyruvate and transfer of an acetly to a CoA
What is the structure of pyruvate dehydrogenase
It made up of the 3 enzymes: Thiamine pyrophosphate as its prosthetic group + Dihydryolipoil transacetylase that use limoaide + CoA as prosthetic group and finally Dihydrolipyl dehydrogenase that use FAD and NAD+ as cofactors
Whar are enzyme complex for
To speed up reactions due to the product is made for 1 reaction that is then quickly passed to the next enzyme to be used in another reaction without leaving the enzyme complex
The product of the reaction may very reactive and take part in unwanted side reaction if not directly tranferred to the next enzyme
What is the TCA cycle
There are 8 main steps and in total there are 3 NADH, 1 FADH2 , 1ATP generatred with 4 REDOX steps
What is Step 1 of the TCA cycle
This is where acetyl CoA is fed into the TCA by being added to oxaloacetate to produce a citrate molecule
This is done by the enzyme citrate synthase and is a condensation reaction so a wayer molecule is removed
Is a Synthase reaction
What is step 2 of the TCA cycle
This is where citrate is converted to isocitrate by an isomerism reaction as convert tertiary alcohol to the secondary alcohol due to being more reactive
Done by the enzyme Aconitase that has a Fe-S co factor
Is an isomerisation reaction
What is step 3 of the TCA cycle
Where Isocitrate is converted to oxalosuccinate that is then converted to a- Ketoglutarate, this is done by the same enzyme called isocirate dehydrognease
The conversion of isocitrate to oxalosuccinate is a Redox so then NAD+ is reduced to NADH that then go to the ETC
Is a oxidative decarboxylation reaction
What is step 4 of the TCA cycle
This is where a-ketoglutarate is converted to succinly CoA
This is the 2nd redox step due to NADH is created and the enzyme used is a-ketoglutarate dehydrogenase
Is a oxidative decarboxylation reaction
What is step 5 of the TCA cycle
Succinly CoA is converyed to succinate by the enzyme succinly CoA synthetase
There is the creation of a GTP that is driven by the hydrolysis of thioether bond to give energy to form the phossphoanhidride bond of GTP and so get substrate level phosphorylation
What is step 6 of the TCA cycle
Succinate is converted to fumarate by succinate dehydrogenase and FADH2 is produced from FAD+
The enzyme used is an integral membrane protein in the krebs cycle and the onlty krebs cycle enzyme in the inner membrane and the active site facing the matrix
Is oxidative dehydrogenation
What is step 7 of the TCA cycle
This is where fumarate is converted to Malate by fumarase and water is added to make an OH group to then give malate
Os stereospecific trans additon reaction
What is step 8 of the TCA cycle
Malate is converted back to oxaloacetate to restart the cycleby the enzyme malate dehydrogenase and NADH is produced from NAD+ reduction
Due to a + delta G the reaction doesnt want to happen but is driven by the very low concentrations of the oxaloacetate in the mitochondria
This is oxidative dehydrogenation
What is the energy balance for complete catabolsim of Pyruvate
3 NADH , 1 FADH2 , 1GTP
This is equal to 12 ATP + 2 ATP + 1 ATP = 15ATP
Due to 2x pyruvate per cycle = 30ATP in total produced
How is NADH transported into the mitochondria
NADH cant be directly moved into the mitochondria due to specific NADH transporters in the membrane but there are porins to allow NADH access to the intermembrane space
It is moved by the Malate - Asparate shuttle
How does the malate - asparate shuttle work
There are 2 specific mitochondrial transporters malate and asparate that lead to electron and proton from the cytoplasmic nADH being transferred to NAD+ in the matrix that can then to produce malate from asparate with oxaloacetate as an intermediate that can then pass into the matrix, in the matrix the malate is converted back to oxaloacetate and then asparate, and this then reproduce the NADH in the matrix and then asparate is moved back out the mitochondria so then cycle can restart
What is the glycerol phosphate shuttle used for
used for metabolically active tissue such as neurons and muscles
There are 2 glycerol 3 phosphate dehydrogenase enzymes that have their active site on the intermembrane side
The glycerol 3 phosphate is made in the cytoplasm and then moved to the intermembrane space
DHAP then cann pass into the cytoplasm to be used in glycolysis and there is 1 less ATP made to allow the reaction to be reversible and to decouople matrix NADH from mitochondria NADH
What is oxidative phosporylation
Glycolysis has a net gain of 2 ATP, 2NADH andf the TCA cycle has 1 ATP, 4NADH , 1FADH2 made
The generation of ATP from the oxidation of coenzymes lead to the phosphorylation of ADP
It happens in the mitochondria
What is the electron transport chain
NADH and FADH2 pass their electrons to the transmembrane protein complexes in the inner membrane
At each step the electrons are passed to the next carrier and each one has a greater electron affinity that previous one
At the end the final step the e- are passed to oxygen to make a water molecule as oxygen is final electron acceptor
Both oxidations are exergonic so are sponatenous of pulling electrons away from the reduced form to the go to the oxygen and the energy of the oxidation drives 3 ATP for NADH and 2 ATP for FADH2
How is the ETC coupled to the ATP synthesis
Energy generated as electrons move down the chain are passed from the coenzymes to the oxygen and then the energy goes to then pump H+ out of the matrix into the intermembrane space across the inner membrane to create a proton gradient across the impermeable inner membrane
This formation of proton gradient and electrical gradient lead to ATP synthesis due to H+ back to matrix via ATP synthase transporter that will move H+ and also phosphorykate ADP to ATP
What is the structure of the H+ pump
Thers are low H+ conc in the matrix and high in the intermembrane space
There are 2 gates and both cant be open at the same time
The H+ bind to the gate and open it in matrix side due to the high binding affinity that H+ has with negative charged amino acid chains.
At the other gate there is a conformational change and opening the gate to push the H+ out to the intermembrane space due to lowering the affinity in the gate so H+ is released
What complexes are used to oxdisise NADH
Complex 1, 3, 4
What happens in complex 1 to oxidise NADH
NADH ubiquinone reductase is the complex and it will oxidise the NADH and trafer 2 e- to the ubiquinone that is accompanied by 4 protons to move into the intermembrane space
There is then reduced ubiquinone that can dliver 2 e- to complex 3
What does complex 3 do for NADH oxidation
Transfer 2 e- between 2 mobile carriers and will get the electrons from the ubiquinin in complex 1
There is 1 e- moved to the cytochrome C woth a 2H+ transfer and 2nd e- is stored internally and then moved to another cytochrom C with 2 more H+ going to the intermembrane space
So overall 4H+ are moved to the intermembrane space from complex 3
What does Complex 4 do
Tranfer 2 e- from 2 cytochrome C to the matrix along with pumoing 2 H+ to the intermembrane space
The 2e- then will reduce oxygen to form a water molecule
Balance sheet of NADH oxidation
Lead to 10H+ being tranfered to the intermembrane space to create the proton gradient
How is FADH2 oxidised
This takes place in complex 2 where succinate is converted to fumarate and FAD+ is reduced
There is no use for complex 1 but will then still use complex 3,4 as well so get a total H+ transfer of 6H+
Why is the proton gradient generated
The proton gradient is used to drive ATP synthesis anf there are 3 H+needed for the creation of 1 ATP molecule
What is the structure of ATP synthase
It is made of a static part as well as a motor region that move all the time by rotating due to movement of protons back to the matrix
The central stalk has a protrusion and this rotate around alpha and beta domains that make up the ATPase head in the stattic part
What do different subunits in the static head of ATP synthase do
Alpha subunit bind to the ATP but not used in the production of ATP
Beta subunit bind to tje ADP + pi and will rotate in 3 states around to make ATP
Loose state - Bind ADP + Pi tightly
Tight state - Bind ATP more tighly so that favour ATP production from ADP+Pi
Open state - Bind ADP or ATP loosely to allow their dissociation
How is rotation the driving force for ATP synthesis
The return of the H+ cause the roattion of the spindle of the ATP synthase
The rotor is asymmetric and cause a conformational shape change in the head group of the units in the static part and the changes drive the ADP + Pi --> ATP
Why is ATP made on demand
Due to not being able to be stored due to unstable
ATP is the fuel in a cell but not between cells
Glucose is used to transfer energy between cells and ATP only made when needed
The inner membrane is highly impermeable so then specific transporters are needed to make ATP on demand due to being made in the matrix so then has to leave the mitochondria
There is an ATP/ADP transporter that have to have both molecules moving through in opposite directions
It is driven by the voltage gradient due to -4 charge on ATP prefer to move pout whilst -3 charge of ADP wants to move in
If there is no ADP then the ATP cannot leave the mitochondria this may be caused due to lack of energy demand so then low ATP hydrolysis to make ADP
Also no ADP will then lead to H+ being unable to return to mitochondrila mtrix as not enough ADP to bind to Beta units