1
Q
What is a metabolic pathway?
A
Series of enzyme-catalyzed reactions
- metabolites or metabolic intermediates are the chemical intermediates
2
Q
The chemical intermediates of metabolic pathways are called:
A
Metabolites or Metabolic intermediates
3
Q
All metabolic pathways share the same ________ and _________ principles
A
All metabolic pathways share the same fundamental chemical and thermodynamic principles
4
Q
What are the two major purposes of Metabolism?
A
- To obtain usable chemical energy from the environment
- capturing solar energy (photosynthesis)
- Consuming and breaking down nutrient molecules
- To make the specific molecules that cells need to live and grow
5
Q
Differences between cells in different organisms and different tissues come down to differences in their ________
A
Differences between cells in different organisms and different tissues come down to differences in their metabolic pathways
- active sets of pathways determine function
6
Q
What are anabolic pathways?
A
Pathways that use energy (require energy) to build larger molecules and are generally reductive (electrons are used to make new bonds)
- Biosynthesis
- Require precursors
7
Q
What are catabolic pathways?
A
Release energy (some of which is stored) and are generally oxidative (electrons are removed as bonds are broken)
- generates precursors (building blocks)
8
Q
What are amphibolic pathways?
A
Operate in both catabolic and anabolic processes (depending on conditions)
9
Q
Three ways food can be used in metabolism?
A
- Generate oxidized carbon atoms (CO2) and water = catabolism
- Generate metabolic intermediates for formation of cellular constituents (Anabolism)
- Storage of energy as ATP (electrons as reduced cofactors for anabolism)
10
Q
What are the four dietary macromolecules?
What do they break down into?
A
- Nucleic acids
- nucleotides
- not a significant fuel source
- Proteins
- amino acids
- Polysaccharides (complex carbs)
- monosaccharides (simple sugars)
- Triacylglycerol (Fat)
- fatty acids
11
Q
Which two dietary macromolecules are the most significant fuel sources (ie greatest energy)?
A
Polysaccharides and Triacylglycerol
12
Q
Are proteins used as an energy source?
A
Not typically. We can, but the amino acids are better used elsewhere
- We use the amino acids from protein breakdown for the synthesis of our own proteins
- Some amino acids we can only get from outside sources
13
Q
Between polysaccharides and triacylglycerol, which provides more energy?
A
Triacylglycerol (fat) provides more energy than polysaccharides (carbohydrates) but both represent significant fuel sources.
Both are less useful as precursors than they are as energy sources
14
Q
What is something that we ALWAYS need?
A
ATP/energy
Food is catabolyzed into ATP
15
Q
What happens in the absence of food?
A
Fuel stores are mobilized to generate fuel molecules which can be catabolized to form ATP
16
Q
The way we store fuel molecules depends on ________
A
The way we store fuel molecules depends on the type of molecules in question
- Major ways:
- As carbohydrates
- As fats
17
Q
Carbohydrates are generally stored as _________ in the _________ and ________
A
Carbohydrates are generally stored as glycogen (polymer of glucose molecules) in the liver (hepatocytes) and skeletal muscle (myocytes)
18
Q
Fatty acids are stored as ______ in _________
A
Fatty acids are stored as fat (triacylglycerols) in adipocytes
19
Q
Liver glycogen is used to maintain:
Skeletal muscle glycogen is used for:
A
Liver glycogen is used to maintain a Continuous glucose supply in the bloodstream.
Skeletal muscle glycogen is used for those myocytes = not available for the rest of the body
20
Q
Which stores more energy, fat or carbohydrates?
A
Fat (1g:7g ratio)
Fat can store the same amount of energy in one gram as carbohydrates can in 7g
21
Q
What is the biochemical standard state?
(what the prime ‘ refers to in deltaG‘)
A
- pH 7 ([H+] = 10^-7 M)
- (for chemical standard state, [H+] = 1 M)
- [Substrates] & [Products] = 1M
- consistent with chemical standard state
- Temperature = 25°C/298K
- Pressure = 1atm
- [H2O] = 55M
- (large amount of water)
IGNORE [H+] and [H2O] in calculations
22
Q
A reaction will only proceed in the forward direction when the associated value of deltaGreaction is _______
A
A reaction will only proceed in the forward direction when the associated value of deltaGreaction is negative (<0)
- ACTUAL delta G (not the standard delta G)
- ΔG’reaction = biochemical actual
- ΔG’°reaction = biochemical standard
23
Q
What is the difference between:
ΔG’reaction
ΔG’°reaction
A
ΔG’reaction = biochemical actual free energy change
ΔG’°reaction = biochemical standard free energy change
24
Q
How would you describe a reaction with
ΔG’reaction < 0
A
ΔG’reaction < 0
Reaction is:
- exergonic
- spontaneous
- “down-hill”
- Favourable
- Reaction proceeds forward
25
What will the reaction look like for each of the following free energies?
* ΔG'reaction \> 0
* ΔG'reaction \< 0
* ΔG'reaction \<\< 0
* ΔG'reaction ~ 0
* ΔG'reaction \> 0
* Reaction will not occur in the forward direction
* ΔG'reaction \< 0
* Reaction will occur (spontaneous)
* ΔG'reaction \<\< 0
* Reaction may be considered irreversible (A → B)
* if reaction is always negative under physiological conditions
* ΔG'reaction ~ 0
* Reaction is considered reversible
* Close to Equilibrium (B←→C)
26
A metabolic pathway is a series of __________ and each individual rxn must obey \_\_\_\_\_\_\_\_\_\_
A metabolic pathway is a series of _enzyme-catalyzed chemical reactions_ and each individual rxn must obey _thermodynamic laws_
* Free energy change must be negative (ΔG \<0)
* ΔG = ΔH -TΔS
* ΔG = ΔG°' + RTln( [products] / [reactants] )
* **the net free energy change for the pathway must be negative for a pathway to proceed**
27
True or false:
Both catabolic and anabolic pathways require a Net ΔG\<0 for a pathway to proceed
True
28
Metabolic pathways exist in a ________ (concentrations of metabolic intermediates often don't change significantly)
Metabolic pathways exist in a _steady-state_
* Water example: as long as water coming in is equal to water out, the pool stays at constant level (steady state)
29
Which steps of a metabolic pathway are typically regulated?
* **Irreversible** steps are usually regulated
* Reversible steps are not usually regulated
30
What is the "rate-limiting step"?
The rate-limiting step in a pathway is the irreversible, regulated reaction that determines the overall rate of the pathway
31
Which enzymes in the image would you expect to be regulated?
Enzymes E1 and E4 are likely to be regulated
32
What is product inhibition?
When our enzyme is inhibited by the immediate product of its reaction
33
What is feedback inhibition?
An enzyme is inhibited by a metabolite further down the pathway
34
What is the image depicting?
Activation of an enzyme by a metabolite "upstream"
35
What does feed-forward activation ensure?
Activation ensures that the pathway is functioning in concert (otherwise intermediates may accumulate)
"feed-forward activation"
(Allosteric patterns of activation)
36
Feed-forward activation and Feed-back inhibition are examples of ________ regulation.
Feed-forward activation and Feed-back inhibition are examples of _allosteric_ regulation
* Feed-forward activation = positive heteroallostery
* Feedback inhibition = negative heteroallostery
37
Opposing pathways catalyze the "reverse" of another pathway, but what has to happen if there are any irreversible steps?
* the irreversible reactions must be replaced or bypassed
* eg Using ATP to reverse the reaction
38
What is reciprocal regulation?
Pathways are regulated to ensure that both do not operate simultaneously.
Ie allows us to have simultaneous control over both pathways (something that might activate E1 and inactive E6)
* eg liver can reversibly convert glucose to pyruvate but if both pathways are operating simultaneously, we would be wasting energy
* ATP hydrolysis would occur but nothing would be achieved
39
How might reciprocal regulation be achieved?
* Enzyme regulators
* Phosphorylation
There are different patterns of regulation
40
What is a high-energy intermediate?
A molecule that contains usable energy of some kind
Ie Compounds which contain "usable" chemical energy
* Energy can be recovered or used
* Simple reaction associated with large delta G (\>20kJ/mol released)
41
What are the three major types of high-energy intermediates?
1. Electron carriers (NADH, NADPH, FADH2, FMNH2) (**reduced cofactors**)
* NAD+, NADP+, FAD, FMN are electron acceptors (oxidized forms)
2. Nucleotide triphosphates (NTPs: ATP, UTP, GTP)
3. Thioesters
42
Nucleotide triphosphates have a large free energy change associated with _______ of the ________ bond
Nucleotide triphosphates (ATP, GTP) have a large free energy change associated with _hydrolysis_ of the _phosphoanhydride_ bond
43
How is a thioester formed?
Thioester = looks like an ester but has a Sulfur as part of the ether linkage
Formed from **condensation** of **carboxylic acid** and **Thiol**
44
Hydrolysis of a thioester would yield:
(Energy change?)
A carboxylate and a thiol (HS-R'')
* delta G°' ~ 30kJ/mol (comparable to ATP hydrolysis)
45
Catabolic processes are _________ (oxidative or reductive)
Catabolic processes are _Oxidative_ (oxidative or reductive)
* Metabolites are oxidized (lose electrons)
* Cofactors are reduced
* Typically **NAD+** (oxidation of C-O bonds), **FAD** (C-C bonds)
* become reduced
* Accept electrons
46
Anabolic process are _________ (reductive or oxidative?)
Anabolic processes are _reductive_
* Metabolites are **reduced** (gain electrons)
* Cofactors are oxidized (reducing agents - becoming oxidized)
* Typically NADPH (electron carrier)
* NADH/FADH2 also involved in reduction reactions
* Reduced cofactors become oxidized during reaction
47
Nucleotides play a central role in metabolism as electron \_\_\_\_\_\_\_
Nucleotides play a central role in metabolism as *electron* _carriers_
* NAD+
* Nicotinamide adenine dinucleotide
* FAD
* Flavin adenine dinucleotide
* The **nitrogen** base portion of these dinucleotides enables them to undergo a reversible reduction reaction
48
What characteristic of NAD+ and FAD allows them to undergo a reversible reduction reaction?
The nitrogen base portion (Non-adenine) (nicotinamide or flavin rings)
49
What can we determine about NAD+ based on the image?
NAD+ is a dinucleotide
* base, sugar, phosphate joined together by a **phosphoanhydride bond**
* No directionality because it's a 5' to 5' linkage (unlike 5' -\> 3' in nucleic acids)
50
What is the difference between NAD+/NADH and NADP+/NADPH
Whether or not there is a phosphate attached to the 2' position on the adenosine nucleotide portion of the molecule
51
What is the Net charge of NAD+ (NADP+)
* Nicotinamide base carries a positive charge
* both phosphate groups carry negative charge
* Net Negative charge
* NAD**+** (+ = oxidized)
52
What is a difference between the reduction of FAD and the reduction of NAD+
FAD takes up 2H+ and 2e- to become reduced
NAD+ takes up 1H+ and 2e- to become reduced
53
What is the molecule in the image?
FAD
54
NAD+ and NADP+ are typically _______ while FAD is a _______ group
NAD+ and NADP+ are typically _cosubstrates_ while FAD is a _prosthetic_ group
55
Implications of the statement: NAD+ and NADP+ are typically cosubstrates while FAD is a prosthetic group
* NAD+ being a cosubstrate means it can be reduced NADH and then dissociate from enzyme active site and if a reaction is to proceed in oxidative direction (catabolic pathway) then we can't just let the [NADH] accumulate there has to be another process to oxidize that cofactor back to NAD+
* For a cosubstrate that means it dissociates and then oxidation happens
* On the other hand, FAD being a Prosthetic group has to be oxidized back to original state in order for enzyme to continue normal functioning
56
What is implied about the function of FAD/FADH2 as a prosthetic group?
As a prosthetic group, FADH2 must be reoxidized back to FAD for the next enzyme cycle to occur.
* eg in the citric acid cycle, coenzyme Q is used to carry out this reoxidation
57
FAD is often involved in ______ bond oxidation
C-C
(single to double)
58
ATP is a high energy compound because of _____________ bonds
ATP is a high energy compound because of _phosphoanhydride_ bonds (2)
59
What results if you hydrolyze one of the phosphoanhydride bonds in ATP?
There is a large negative free energy change associated with the hydrolyses process
* delta G (ΔG'°) ~-32 kJ/mol for the hydrolysis of a phosphoanydride bond
* ΔG'° ~ +32 kJ/mol for the formation of a phosphoanydride bond
60
What is the free energy change for the formation and hydrolysis of a phosphoanhydride bond?
* delta G (ΔG'°) ~-32 kJ/mol for the hydrolysis of a phosphoanydride bond
* ΔG'° ~ +32 kJ/mol for the formation of a phosphoanydride bond (requires energy)
61
What is it about ATP (phosphoanydride bonds) that make it high energy?
1. Decreased electrostatic repulsion
* ATP has a high number of negative charges focussed near each other
* Hydrolysis reduces the number of negative charges
2. Resonance stabilization
* lowers free energy associated with formation of that particular molecule
3. Solvation effects
* one big molecule -\> two smaller molecules = entropically more favourable (can better associate with water)
62
Compare resonance stabilization of Terminal phosphoryl group of ATP and Inorganic Phosphate (Pi)
* Terminal phosphoryl group of ATP
* 3 resonance structures
* Inorganic Phosphate
* 4 resonance structures
* 4Oxygens, all of which are roughly equivalent to eachother
* proton (H+) weakly associated with phosphate group = not formally attached to any Oxygen
63
What is a Thioester?
* High energy compound
* Similar to esters but with no e- delocalization
* increased energy of substrates
* hydrolysis of thioester is associated with Same kind of energy change as hydrolysis of phosphoanhydride bond (ATP) (delta G'°~30 kj/mol)
64
Why are thioesters high-energy compounds while esters are not?
* No delocalization of electrons in thioesters
* substrates in thioester have a *higher energy* than the substrates in esters
* Because energy starts higher, the free energy change (delta G) is MORE NEGATIVE for thioesters (see graph in image)
65
ATP is generated by __________ (catabolism or anabolism)
ATP is generated by _catabolism_
66
ATP is generated by catabolism:
May be generated by which two means?
ATP is generated by catabolism:
May be generated
* directly: _"substrate-level phosphorylation"_
* Direct product of a chemical reaction
* Oxidative Phosphorylation
* Via reoxidation of NADH/FADH2
67
What are three ways ATP might be used?
* Driving unfavourable reactions (coupling)
* coupling = reactions that require input if energy
* Movement (muscle, flagella)
* hydrolysis of ATP
* chemical energy into kinetic energy
* Primary Active Transport
* Variation of coupling
* Taking an unfavourable process and using energy to drive it
68
Analyze the graph
* Graph illustrates the negative free energy change associated with the hydrolysis of ATP (delta G'° = -32kJ/mol
69
What is the first reaction in glycolysis?
What is the free energy associated with that reaction?
How do we make it proceed in the forward direction under standard conditions?
The **phosphorylation** of glucose to form **glucose-6-Phosphate**
* Free energy:
* If we just add a Pi to glucose, the free energy associated with the reaction is positive 14kJ/mol (delta G \> 0 = unfavourable)
* won't proceed in forward direction under standard conditions
* If we add ATP to glucose the associated free energy change is the sum of the two reactions (-32 + 14 = -18kj/mol)
* delta G \< 0 is favourable = will proceed in forward direction
* Coupling synthesis of glucose-6-phosphate to the hydrolysis of ATP
* no actual hydrolysis as the Pi from ATP is being directly transferred to glucose
70
What is the free energy associated with the phosphorylation of glucose to form glucose-6-phosphate in glycolysis? Will this reaction proceed in the forward direction under standard conditions?
* Free energy:
* If we simply add a Pi to glucose, the free energy associated with that reaction is positive **14kJ/mol** (delta G \> 0 = unfavourable)
* won't proceed in the forward direction under standard conditions
* If we add ATP to glucose the associated free energy change is the sum of the two reactions (**-32 + 14 = -18kj/mol**)
* delta G \< 0 is favourable = will proceed in forward direction
* Coupling synthesis of glucose-6-phosphate to the hydrolysis of ATP
* no actual hydrolysis as the Pi from ATP is being directly transferred to glucose
71
How is ATP used in the synthesis of glucose-6-phosphate during glycolysis?
Coupling synthesis of glucose-6-phosphate to the hydrolysis of ATP
* no actual hydrolysis as the Pi from ATP is being directly transferred to glucose
72
Free energy changes for reactions are \_\_\_\_\_\_\_. Combined reactions must have a NET free energy change _____ than 0 to be spontaneous.
Free energy changes for reactions are _additive_. Combined reactions must have a NET free energy change _less_ than 0 (ΔG'°
73
What is "Phosphate transfer potential"?
Phosphate-transfer potential refers to free energies of hydrolysis for phosphate-containing compounds
74
Name the two glycolysis intermediates given in lecture that have phosphate transfer potential. What are their free energies of hydrolysis?
1. Phosphoenolpyruvate ΔG'° = -62 kj/mol
2. 1,3-*Bis*phosphoglycerate ΔG'° = -49 kj/mol
75
What compound with phosphate transfer potential is a short term supply of ATP in muscle?
Phosphocreatine ΔG'° = -43 kj/mol
76
Acetyl CoA is a ________ with phosphate transfer potential of \_\_\_\_\_\_\_
Acetyl CoA is a _thioester_ with phosphate transfer potential of _ΔG'° = -32 kj/mol_
77
The hydrolysis of phosphocreatine produces:
creatine and Pi
78
The polar head group (functional group) of creatine is derived from __________ and has a partial ______ charge distribution associated with all the Nitrogens because of _\_\_\_\_\_\_\_\_\_\__
The polar head group (functional group) of creatine is derived from _a guanido group_ (The basic-group configuration C = (NH2)2 found in arginine) and has a partial _positive_ charge distribution around all the Nitrogens because of _resonance stabilization associated with that functional group_
79
What is the overall free energy change (ΔG'°) associated with the coupled reaction of phosphocreatine and ADP
Phosphocreatine + ADP ←→ creatine + ATP
Reaction 1: ADP + Pi + H+ -\> _ATP_ + _H20_ = phosphorylation of ATP: ΔG'°= +32
Reaction 2: Phosphocreatine + H2O → creatine + Pi : ΔG'°= -43
Therefore, the NET free energy change = -43 + 32 = -11kj/mol
80
Why is the coupled reaction of phosphocreatine to creatine and ADP to ATP reversible under actual conditions?
* [ATP] and [Phosphocreatine] and [creatine] are **not** going to at **1.0M**
* Phosphocreatine acts as a temporary store of chemical energy
* when ↑[ATP] the reaction favours the formation of phosphocreatine
81
What are four sources of ATP based on time and accessibility?
* ATP on hand:
* Immediately available
* Lasts a couple seconds
* "throw something"
* ATP from Phosphocreatine
* Provides ATP after the on-hand ATP has been used up
* lasts a few seconds
* ATP from Anaerobic processes
* Substrate level phosphorylation processes
* ATP from Aerobic processes
* Generates significant amounts of sustainable ATP over extended period of time
