Health and Disease Week 21 Flashcards
define metabolism
the inter-conversion of biomolecules using chemical reactions
define catabolic
breakdown into smaller products
By which process do catabolic reactions usually work?
oxidation
What are examples of catabolic reactions?
- production of chemical energy (ATP) and ion gradients
- production of mechanical energy
- production of reducing equivalents
- production of biosynthetic precursors
define anabolic
small products reacting to form bigger ones
By which process do anabolic reactions usually work?
reduction
What are examples of anabolic reactions?
- storage of energy by making more complex molecules
- production of macromolecules and cellular structures
What is the equation for Gibbs free energy of any reaction?
∆G= ∆G°+RT ln[products]/[reactants]
What are endothermic reactions often driven by?
ATP or pyrophosphate hydrolysis (removal of products)
By how much can hydrolysis of ATP change the equilibrium constant of a coupled reaction?
by 10^8 - changes a reaction from not very favourable to very favourable
How many stages of glycolysis are there?
3
What is the 1st stage of glycolysis?
activation of glucose
Where does glycolysis occur?
the cytosol
Glycolysis stage 1 step 1
a molecule of ATP is added to glucose to make glucose-6-phosphate to lock glucose inside the cell
glycolysis stage 1 step 2
glucose-6-phosphate is rearranged to form fructose-6-phosphate
glycolysis stage 1 step 3
ATP and an enzyme called phosphofructokinase 1 adds another phosphate group to the opposite end of fructose-6-phosphate, forming fructose-1,6-bisphosphate
glycolysis stage 1 step 4
the double phosphorylation causes the ring opening of the sugar and it becomes activated
glycolysis stage 2 step 1
fructose-1,6-phosphate in opened form is converted into dihydroxyacetone phosphate AND glyceraldehyde-3-phosphate by the enzymes aldolase
glycolysis stage 2 step 2
dihydroxyacetone phosphate is converted into another molecule of glyceraldehyde-3-phosphate by the enzyme triose phosphate isomerase
Why do we need to convert dihydroxyacetone phosphate into glyceraldehyde-3-phosphate?
otherwise you would only be able to process 1/2 the glucose molecule
glycolysis stage 3 step 1
glyceraldehyde-3-phosphate is oxidised to 1,3-bisphosphoglycerate using a molecule of phosphate and NAD+ - NAD+ is reduced to NADH
glycolysis stage 3 step 2
ADP is converted to ATP to transfer the phosphate group from 1,3-bisphosphoglycerate to it to form 3-phosphoglycerate
glycolysis stage 3 step 3
phosphate group is moved to form 2-phosphoglycerate
glycolysis stage 3 step 4
dehydration reaction produces a double bond forming phosphoenolpyruvate
glycolysis stage 3 step 5
phosphoenolpyruvate is a high energy intermediate and ADP is used to transfer phosphate group onto it to form ATP and pyruvate is formed
What are the steps of anaerobic respiration?
- in the absence of oxygen, the pyruvate is reduced to R-lactate
- this oxidises NADH to NAD+ and allows glycolysis to continue
- R-lactate can be reoxidised to pyruvate using NAD+ eventually
What is the function of the Cori cycle?
the Cori cycle recycles R-lactate to glucose
Why is it so difficult to use lactate in the muscles?
you need to transport it to the liver which has additional enzymes to recycle it to glucose
What are the steps of the Cori cycle?
- lactate is transported from the muscle to the liver via the blood
- lactate dehydrogenase in the liver converts R-lactate to pyruvate
- pyruvate is converted to glucose by gluconeogenesis
What does the Cori cycle require?
input of 6x ATP per glucose molecule
define gluconeogenesis
the process of converting lactate into pyruvate in the liver
How many pyruvates are required to make each glucose molecule?
2
gluconeogenesis step 1
ATP, CO2 and pyruvate make oxaloacetate in the mitochondria of liver cells
gluconeogenesis step 2
oxaloacetate is exported to the cytosol and converted into phosphoenolpyruvate - 2 x ATP and 1 x NADH are needed for this step
Why does gluconeogenesis go via the oxaloacetate intermediate?
there is a big energy barrier from pyruvate to phosphoenolpyruvate - oxaloacetate makes it smaller
gluconeogenesis step 3
additional enzymes are also needed where steps in glycolysis are irreversible - however, the steps are identical but in reverse
Why is the Cori cycle needed?
because additional enzymes are needed to reverse pyruvate back to glucose
What does the pyruvate dehydrogenase complex control?
the entry of pyruvate into the TCA cycle
How many subunits does the pyruvate dehydrogenase complex have?
3
What are the 3 subunits of the pyruvate dehydrogenase complex?
E1, E2, E3
What is E1?
pyruvate dehydrogenase
What does E1 do?
decarboxylates pyruvate
What else does decarboxylation of pyruvate need?
TPP
What is E2?
dihydrolipoyl transferase
What does E2 do?
makes CoA - also requires lipoamide
What is E3?
dihydrolipoyl dehydrogenase
What does E3 do?
converts reduced lipoamide to its disulfide form - also required FAD
What are the steps for the formation of acetyl-CoA?
- TPP anion adds to pyruvate and CO2 is released, forming a product with an -OH group
- lipoamide disulfide is added to the acetyl group and a redox reaction occurs
- disulfide exchange occurs to form acetyl-CoA and reduced lipoamide
- reduced lipoamide is oxidised to its disulfide form using FAD
- FADH2 is oxidised by NADH, which is fed into the electron transport system
How many main phases does the tricarboxylic acid cycle have (TCA)?
4
What are the main phases of the TCA?
- condensation and rearrangement
- decarboxylation
- formation of GTP using phosphate anhydride bond
- conversion of succinate to oxaloacetate
TCA cycle phase 1
- citrate synthase catalyses a condensation reaction of acetyl-CoA and oxaloacetate
- hydrolysis of the CoA ester makes this reaction irreversible
- citrate undergoes a rearrangement to form isocitrate - preparing for stage 2
TCA cycle phase 2
- isocitrate is converted to a beta-ketoacid using NAD+ forming NADH
- the beta-ketoacid spontaneously loses CO2, forming 2-oxoglutarate
- 2-oxoglutarate is converted to succinyl-CoA by a multi-enzyme complex
- the reaction is the same as that carried out by pyruvate dehydrogenase - the same cosubstrates are needed (TPP, lipoamide, CoA-SH)
TCA cycle phase 3
- CoA is displaced by inorganic phosphate to produce a mixed acid anhydride
- phosphate is tranferred to an active site on histidine
- the phoshate is then transferred off the enzyme onto GDP, forming GTP
TCA phase 4 - this stage converts succinate into oxaloacetate
- desaturation of the C-C bond using FAD occurs - inserts a double bond
- hydration of the double bond to form S-malate
- NAD+ is used to oxidise S-malate to oxaloacetate
Where does oxidative phosphorylation occur?
in the mitochondria
How are electrons transferred in oxidative phosphorylation?
from NADH and FADH2 via series of electrons donor to oxygen
What is the function of the many proteins inserted into the mitochondrial membrane?
to pump protons from the inside of the mitochondria into the intermembrane space
How much of the ATP in respiration does oxidative phosphorylation produce?
about 26 of 30 molecules of ATP from the COMPLETE oxidation of glucose
What are the 2 main parts of oxidative phosphorylation?
- generation of a proton gradient
- production of ATP
What is the value of the potential difference that exists between NADH and O2?
1.14 V which corresponds to 220KJ mol -1 of energy - this is sufficient energy to make several molecules of ATP
What do many of the proteins (enzymes) in the mitochondrial membrane contain? Why?
metal centres e.g. haem that are usually redox active - so you can do oxidation and reduction
How big is the proton gradient that the voltage difference produces?
1.4 pH units - pH difference of over 10x
Which 2 steps does ATP synthesis consist of?
- proton transport into the mitochondrial intermembrane space
- proton gradient transports protons through the inner membrane by ATP synthase and uses the energy provided to make ATP
What is the Mitchell hypothesis 1961?
the amount of ATP synthesis is coupled to the proton gradient
Is the synthesis of ATP from ADP and Pi spontaneous?
yes
What is the proton gradient actually needed for if the synthesis of ATP is spontaneous?
driving the RELEASE of ATP from the enzyme to allow binding of more ADP and inorganic phosphate
Where is the energy from that is used to transport protons across the inner mitochondrial membrane?
the electron transport system
What are the products of glycolysis?
2 x NADH (cytosolic) - NADH produced in the cytosol yields less ATP as energy is expended importing it into the mitochondria
2 X ATP
What is the ATP yield from glycolysis?
5
3 from NADH, 2 from ATP
Why does NADH production in glycolysis produce 3 molecules of ATP?
the NADH can enter the electron transport chain, where each molecule of NADH can produce approximately 2.5 molecules of ATP
What are the products from the oxidation of pyruvate to acetyl-CoA?
2 x NADH
What is the ATP yield from the oxidation of pyruvate to acetyl-CoA?
5 ATP
What are the products from the oxidation of acetyl-CoA?
6 x NADH
2 x FADH2
2 x GTP
What is the ATP yield from the oxidation of acetyl-CoA?
20 - 15 + 3 + 2
When is glycogen metabolised?
when energy is needed in the form of glucose, glycogen is broken down to release glucose molecules
Where does glycogen metabolism occur?
in the liver cytosol
What are the steps of glycogen biosynthesis?
- start with glucose
- intermediate glucose-6-phosphate is formed
- isomerisation of glucose-6-phosphate to glucose-1-phosphate by removal then addition of a phosphate group
- UTP is used as an energy source to activate glucose and the last two phosphate groups (pyrophosphate) is broken off
- the rest of the UDP adds to glucose-1-phosphate
- UDP-glucose is an activated form of glucose
- the UDP acts as a recognition site and leaving group for the enzyme that extends the glycogen chain
- now another glucose residue can be added onto it to add to the glycogen chain
What does alpha mean in molecules?
the bond is pointing down
What are the steps of the synthesis of glycogen alpha-1,4 bonds?
- the lone pair on the oxygen in the glucose molecule forms a double bond
- UDP leaves to give an oxycation intermediate
- the oxycation is an electrophile
- the oxycation therefore attacks the lone pair on the -OH of the carbon 4 on another glucose molecule in the glycogen chain
- the lone pair is used to add to the double bond
- a proton from the -OH group is lost
- glycosidic bond is formed
What are the 2 important features of alpha-1,4 glycogen bond synthesis?
- we have retention of stereochemistry!
- the glycogen polymer is extended by 1 glucose molecule
Why is glycogen highly branched?
- increases its solubility
- to allow enzymes to access glucose molecules easily when they are required
How are glycogen alpha-1,6 bonds formed?
a branching enzyme transfers several residues that are linked by alpha-1,4 bonds to the carbon 6 of a glycogen molecule - the chains can be extended and branched further
What are the steps of degradation of glycogen alpha-1,4 bonds?
- 1,4 bonds are degradaed by glycogen phosphorylase
- a proton is used to break the bond
- this addition of a proton forms an oxycation intermediate
- the enzyme adds a phosphate to give glucose-1-phosphate
- this phosphorylated status ensures the glucose cannot move out of the cell and we don’t need to use ATP to activate glucose as phosphorylated residues are active already
- glucose-1-phosphate is converted to glucose-6-phosphate
What is the issue with glycogen phosphorylase?
it cannot break glycogen 1,6 bonds
What are the steps of degradation of glycogen alpha-1,6 bonds?
- glycogen phosphorylase breaks down the linear chain of a branch until we only have 3 alpha-1.4 residues left next to the branch point
- a glycosyl transferase enzyme moves these 3 residues onto the end of another linear chain - effectively just extends the linear chain on a different strand
- the remaining 1,6-linked residue is removed by a 1,6-glycosidase enzyme to produce glucose
- the glucose product is either exported to the bloodstream or converted to glucose-6-phosphate - it can now be used in glycolysis
- a small proportion of the glucose needs to be reactivated during degradation of glycogen
How are fats stored in adipose tissue?
as triacyl-glycerides (triglycerides)
What are triglycerides?
long-term high energy storage molecules that are anhydrous and highly reduced
What are triglycerides hydrolysed into? How?
fatty acids and glycerol - using 3 water molecules and a lipase enzyme
Where does fatty acid beta-oxidation occur?
in the mitochondria
Where is the acyl-CoA made?
in the cytosol from the fatty acid and then imported into the mitochondria by a transporter
Why is acyl-CoA formation only needed in the FIRST round of beta-oxidation?
in the next rounds, the product of the beta-oxidation pathway is the acyl-CoA ester
How many steps and intermediates does beta-oxidation have?
4
When does beta-oxidation stop?
when we have broken down all of the fatty acid into acetyl-CoA
Where does acetyl-CoA go?
it is oxidised by the TCA cycle
What is the 2nd product of beta-oxidation?
the n-2 fatty acyl-CoA
What happens to the n-2 fatty acyl-CoA?
it is further beta-oxidised until it is completely broken down to acetyl-CoA
beta-oxidation step 1
start with a fatty acid
beta-oxidation step 2
ATP used to form the CoA ester - ATP goes to AMP and PPi
beta-oxidation step 3
PPi can by hydrolysed to make this reaction irreversible
beta-oxidation step 4
the adenosine monophosphate produced can react with ATP to make 2 molecules of ATP
beta-oxidation step 5
the acyl-CoA is oxidised by an oxidase enzyme, creating a double bond - this converts FAD to FADH
beta-oxidation step 6
a 2nd enzyme adds a molecule of water to the double bond to form a hydroxy compound - this is stereoselective
beta-oxidation step 7
the alcohol group is oxidised to its corresponding ketone and NAD+ is reduced to NADH
beta-oxidation step 8
another molecule of CoA is used to cleave the bond between the keto group and thioester group
beta-oxidation step 9
when this is cleaved, we are releasing acetyl-CoA and forming the n-2 acyl-CoA
How much ATP does each beta-oxidation cycle yield?
14 molecules of ATP
What does fatty acid biosynthesis require?
acetyl-CoA and CO2
What is needed to make malonyl-CoA in fatty acid biosynthesis?
carboxybiotin and acetyl-CoA
What is carboxybiotin?
a carboxylating agent - important as it controls whether we break down or make fatty acids
Which is the key regulatory enzyme in fatty acid biosynthesis?
acetyl-CoA carboxylase
What is needed to make carboxybiotin?
ATP and CO2 - therefore need to put energy in
What else does fatty acid biosynthesis require?
NADPH (reductant) and other biosynthetic enzyme in the cytosol
What are the steps of formation of malonyl-CoA?
- convert CO2 into bicarbonate using a molecule of water - hydration reaction
- ATP used to donate phosphate group onto bicarbonate, forming carboxyphosphate
- carboxyphosphate is used to generate carboxybiotin
- carboxybiotin (carboxylating agent) is used to convert acetyl-CoA to malonyl-CoA
- when making malonyl-CoA, we replace a hydrogen atom with the CO2 from the carboxybiotin
- we transfer the acetyl-CoA and malonyl-CoA to the acyl-carrier protein for the rest of the biosynthetic pathway
What is the key regulatory step of synthesis of fatty acids?
making malonyl-CoA by acetyl-CoA carboxylase
What is acyl carrier protein (ACP)?
a small protein used to link intermediates in fatty acid biosynthesis
What is the function of acyl carrier protein (ACP)?
the long CoA side chain means the intermediate can be moved from one active site in the fatty acid synthase to the next, so you can efficiently grow the fatty acid chain without losing any intermediate
What are the steps of fatty acid synthesis from malonyl-CoA?
- start with malonyl-ACP and acetyl-ACP
- condensation reaction links them together to form a dicarbonyl compound and CO2 is lost
- the dicarbonyl compound it reduced using NADPH - produces the corresponding hydroxy-ACP derivative
- water is eliminated
- a 2nd molecule of NADPH is used to saturate the double bond to form the saturated ACP-acyl intermediate
- the ACP-acyl intermediate can react with another molecule of malonyl-CoA
- biosynthetic pathway goes round in cycles until about 16 or 18 carbon atoms are reached
