Lecture 2: Bioenergetics in the Cytosol - Exam 3

Question 1

Why is energy important in the cell?

Answer 1

Many tasks that bacteria must perform such as movement and the synthesis of macromolecules, require energy. A large portion of the cell’s activities are therefore devoted to obtaining energy from the environment and using that energy to drive energy requiring reactions.
Many reactions that must take place are energetically unfavorable and are therefore able to proceed only at the cost of additional energy output. The generation and utilization of metabolic energy is thus fundamental to all of cell biology.

Question 2

What are the two different kinds of energy that drive ALL cellular reactions?

Answer 2

Electrochemical energy and high energy molecules in the cytosol (chemical energy).

Question 3

How is electrochemical energy generated?

Answer 3

Ion gradients across the cell membrane (proton gradient: PMF)
- electrochemical energy drives solute transport, flagella rotation, ATP synthesis, and other membrane activities.

Question 4

How is chemical energy (high-energy molecules in the cytosol) generated?

Answer 4

ATP, phosphoenolpyruvate, 1,2-Bisphosophoglycerate, Acetyl-CoA, etc.
- high energy molecules drive biosynthesis in the cytoplasm: synthesis of nucleic acids, proteins, lipids, and polysaccharides.
- some solute uptake is also driven by high-energy molecules rather than electrochemical energy.

Question 5

What is Free Energy (Gibb’s Free Energy)?

Answer 5

Is the energy associated with a chemical reaction that is available (or “free”) to do work.
G = H - TS
(h= enthalpy, t= temperature, s= entropy)

Question 6

What are we really concerned about when talking about Gibb’s free energy?

Answer 6

The change in Gibbs free energy for a process, or delta G.

Question 7

Reactions proceed (spontaneously) only in the direction of..?
If delta G isn’t favorable, the process will not proceed unless..?

Answer 7

Negative delta G ;
energy is expended

Question 8

Biochemical reactions in the cytosol are driven by…?

Answer 8

High energy molecules. These molecules contain a bond with a high free energy of hydrolysis (deltaGprime)… ATP, BPGA, PEP, acetyl-P, acetyl-CoA, succinyl-CoA, etc.

Question 9

While these substances (ATP, BPGA, PEP, acetyl-P, acetyl-CoA, succinyl-CoA) are often referred to as having ‘high-energy’ bonds, free energy of hydrolysis is not..?

Answer 9

Bond energy

Question 10

What is bond energy?

Answer 10

Energy required to break a bond, not energy released when a bond is broken.

Question 11

The standard free energy of hydrolysis can also be thought of as…?

Answer 11

Group Transfer Potential

Question 12

Describe the group transfer potential a little more. Basically, what does it tell you about a molecule?

Answer 12

The group transfer potential is a way to compare the tendency of organic molecules to transfer a chemical group (often a phosphoryl group) to an acceptor molecule.

Question 13

Why are phosphates in biochemical reactions to important?***

Answer 13

Because they are thermodynamically unstable, but kinetically stable (their negative charge resists hydrolysis). This combination of thermodynamic instability and kinetic stability allows the enzyme-catalyzed use of phosphate esters for energy transformation as well as regulation.

Question 14

Describe enzyme catalysts.

Answer 14

Do not require energy, they lower activation energy.

Question 15

How can the cell control the use of phosphate bonds to generate energy?

Answer 15

Energy is stored until the cell needs it. It then produces the enzyme that lowers activation energy and then the reaction can proceed.

Question 16

Molecules with high group transfer potential contain ___________ bonds that can be…?

Answer 16

high energy ;
hydrolyzed to release energy.

Question 17

Molecules with high group transfer potential contain high energy bonds that can be hydrolyzed to release energy. This energy (or transfer potential) can be…?

Answer 17

This energy (or transfer potential) can be transferred to other molecules to form new bonds and linkages.

Question 18

Group transfer potential is the _________ of free energy of hydrolysis. So, here ________ numbers are good.

Answer 18

Negative ;
Positive

Question 19

What molecules are considered to have high group transfer potential?

Answer 19

> 29 kJ

Question 20

How does a reaction with positive delta G of hydrolysis proceed?

Answer 20

By coupling the positive reaction with a reaction with a more negative delta G.

Question 21

Would any of these reactions below proceed spontaneously? How would coupling the two reactions impact reaction? What would the delta G of the coupled reaction be?

ATP + H2O –> ADP + Pi
deltaG’ = -35 kJ/mol
Glucose + Pi –> glucose-6-phosphate + H2O deltaG’ = +14 kJ/mol

Answer 21

The first reaction would proceed spontaneously.
By coupling the two reactions we are able to get a negative delta G. The new deltaG’ is -21 kJ/mol

Question 22

What is ATP?
What is its structure?
Do all the three phosphate groups have the same delta G?

Answer 22

ATP serves as an energy source as the “energy currency of the cell.”
ATP structure: three phosphates with two phosphoanhydride bonds. The three phosphates are connected to the 5’ carbon of a ribose. Adenine is also connected to the 1’ carbon of ribose.
The three phosphates do not have the same delta G.

Question 23

ATP hydrolysis is _______ (endergonic/exergonic), meaning that..?
Why is it endergonic or exergonic?

Answer 23

exergonic, meaning that it releases energy.
The hydrolysis of ATP is exergonic because the triphosphate unit contains two phosphoanhydride bonds that are unstable.

Question 24

Phosphoanhydride bonds have a large _______ deltaG of hydrolysis (deltaG’).

Answer 24

negative

Question 25

Where does the release of energy come from in ATP hydrolysis (and high energy bonds)?

Answer 25

The release of energy from ATP hydrolysis comes from the chemical change to a state of lower free energy, not from the phosphate bonds themselves.

Question 26

ATP can donate…? What does this donation do?

Answer 26

ATP can donate two phosphoryl groups (ATP –> ADP –> AMP), releasing energy with each transfer. The energy released on ATP hydrolysis is used to power a host of cellular functions.

Question 27

When comparing transfer potential of phosphoryl compounds, ATP is ranked where?

Answer 27

Ranked in the middle, yielding significantly less energy than PEP and CP.

Question 28

ATP yields significantly less energy than PEP and CP. What does this allow ATP to do? Why doesn’t the cell use phosphoenolpyruvate instead of ATP then?

Answer 28

This allows ATP to phosphorylate lower-energy compounds, such as G-6-p.
The cell uses ATP because it is easy to make and doesn’t require as much energy to make and the cell still gets a good bit of energy (my own words)
Slides said: It is more difficult to synthesize higher ranking molecules (PEP and CP), and lower ranking compounds don’t release enough energy to perform significant work.
** This makes ATP’s intermediate ranking ideal for its function as the energy currency of the cell.

Question 29

What can ATP be synthesized from?

Answer 29

ADP by PEP and CP phosphorylation

Question 30

Describe the Release of energy by ATP hydrolysis. Which phosphate on ATP has the most energetically favorable deltaG of hydrolysis?

Answer 30

Potentially, 2 ~P bonds can be cleaved, as 2 phosphates can be released by hydrolysis of ATP. The gamma phosphate (at the end) has the most energetically favorable deltaG of hydrolysis.

Question 31

ATP often serves as an energy source as the “energy currency of the cell.” Hydrolytic cleavage of one or both of the “high energy” bonds of ATP is coupled to..?

Answer 31

An energy requiring (non-spontaneous or positive deltaG reaction).

Question 32

Is ATP constantly being utilized by the cell?

Answer 32

Yes!

Question 33

What does AMP function as? Explain.

Answer 33

AMP functions as an energy sensor and regulator of metabolism. When ATP production does not keep up with needs, a higher portion of a cell’s adenine nucleotide pool is AMP. (Basically, the more AMP you have, the less ATP you have, so bonds have been hydrolyzed, and a huge pool of AMP tells the cell that it is low on energy/ATP). AMP stimulates metabolic pathways that produce ATP.

Question 34

ATP has three phosphates, so why isn’t the 3rd (from AMP) not used for energy?

Answer 34

Because not all phosphates are created (energetically) equal.

Question 35

Phosphate esters, like AMP, G-6-P, and G-3-P, aren’t used for energy even though they have a - ΔG°’. Why not?

Answer 35

The linkage between the first phosphate and the ribose hydroxyl of ATP is a phosphate ester. These are formed by splitting out water between a phosphoric acid and an OH group, they have low energy but negative delta G of hydrolysis.

Question 36

What are some other higher energy molecules?

Answer 36

Coenzyme A

Question 37

What bond makes Coenzyme A such a useful high energy molecule?

Answer 37

A thioester forms between a carboxylic acid and the thiol group (SH) of coenzyme A (CoA-SH)

Question 38

Describe Thioesters.

Answer 38

Are high energy linkages. In contrast to phosphate esters, thioesters have a large negative deltaG of hydrolysis.

Question 39

The thiol (-SH) of coenzyme A can react with a carboxyl group (OH) of acetic acid, yielding?

Answer 39

Acetyl-CoA

Question 40

What is essential to the role of coenzyme A as an acyl group carrier?

Answer 40

The spontaneity of thioester cleavage. The spontaneity is a result of the large negative deltaG of hydrolysis.

Question 41

Like ATP, CoA has a ______ group transfer potential.

Answer 41

High

Question 42

Acetyl-CoA participates in?

Answer 42

Many reactions for protein, lipid, and carbohydrate metabolism, including delivering the acetyl group to the citric acid cycle to be oxidized for energy production.

Question 43

Are Acyl derivates of phosphate and Coenzyme A versatile?

Answer 43

Yes!
Depending on the enzyme catalyzing the reaction, they can donate acyl groups, CoA groups or phosphoryl groups.

Question 44

Explain how (3’,5’-cyclic AMP) cAMP is used as a transient signaling molecule.

Answer 44

The lability of cAMP to hydrolysis makes it an excellent transient signal. cAMP binds to a cAMP receptor protein (CRP), which has a DNA-binding domain and a cAMP binding domain. cAMP bound to CRP functions as a transcription factor to activate gene transcription, for example of the lac operon.

Question 45

What catalyzes cAMP synthesis?

Answer 45

The enzyme adenylate cyclase
ATP –> cAMP +Ppi

Question 46

When is cAMP repression used?

Answer 46

cAMP repression is used by cells during glucose-mediated catabolite repression in E. coli.

Question 47

How does adenylate cyclase stimulated?

Answer 47

When no glucose is available, phosphorylated EIIA^GLU stimulates adenylate cyclase to produce cAMP.

Question 48

What enzyme is responsible for regulating cAMP?

Answer 48

EIIA^GLU
It stimulates cAMP production when no glucose is available and ELLA^GLU is phosphorylated. It prevents cAMP production when glucose is available and it is dephosphorylated.

Question 49

When glucose is the carbon source, _________ levels are low. What else would this do?

Answer 49

intracellular cAMP.
Therefore, not much cAMP bound to CRP and no transcription of genes.

Question 50

When glucose levels are low and lactose is present as an alternate energy source, then…?

Answer 50

1. EIIA^glu remains phosphorylated.
2. The phosphorylated EIIA^glu induces adenylate cyclase to produce cAMP.
3. cAMP binds to the cAMP receptor protein (CRP) which activated transcription of the lac operon.
   - Note that in the presence of lactose, lactose binds to the lac repressor which prevents the repressors from blocking transcription of the lac operon.

Question 51

What does the CRP protein look like when glucose levels are sufficient and therefore cAMP levels are low?

Answer 51

The CRP protein has an inactive shape since it is not bound to cAMP and it cannot bind upstream of the lac operon promoter to activate transcription.

Question 52

What is kinetics? What is thermodynamics?

Answer 52

Kinetics describes the rate at which a particular process will occur and the pathway by which it will occur. Laws of movement under force.
Thermodynamics studies interconnection of thermal and other energy forms. Thermo asks the question “Can a reaction take place?” Kinetics asks the question “How fast?”

Question 53

A high activation energy barrier usually causes hydrolysis of a high energy bond to be…?

Answer 53

Very slow in the absence of an enzyme catalyst.

Question 54

Kinetic stability is essential to?

Answer 54

The role of ATP and other compounds with high energy bonds.

Question 55

What would happen if ATP would rapidly hydrolyze in the absence of a catalyst? Basically, how does kinetics connect with ATP?

Answer 55

It could not serve its important roles in energy metabolism and phosphate transfer.

Question 56

When is phosphate removed from ATP?

Answer 56

Only when the reaction is coupled via enzyme catalysis (ATPase, “nucleotide binding domain”) to some other reaction useful to the cell, such as transport of a solute or phosphorylation of glucose.

Question 57

How is ATP synthesis achieved in respiration and photosynthesis?

Answer 57

ATP synthesis is achieved via PMF in respiration and photosynthesis.

Question 58

ATP synthesis via PMF: How is ATP synthesized?

Answer 58

By ATP synthase

Question 59

What is Oxidative Phosphorylation?

Answer 59

Electron transport-dependent phosphorylation
In the case of photosynthesis, photophosphorylation.

Question 60

What is substrate level phosphorylation?

Answer 60

When a phosphate group of a substrate is transferred to ADP to make ATP.

Question 61

What are the two ways that substrate-level phosphorylation can happen?

Answer 61

1) A direct transfer of a phosphate from an organic substrate with a high group transfer potential to ADP. It does not require oxygen, so this a major method for ATP generation during fermentation by anaerobic or facultatively anaerobic bacteria.
2) Formation of a phosphate bond using the energy derived from hydrolysis of a high energy bond. Cells need to first generate compounds with high group transfer potential such as PEP or acetyl-CoA.

Question 62

Explain why phosphoanhydride linkages have a high deltaG hydrolysis.

Answer 62

Factors include:
Resonance stabilization of products of hydrolysis exceeds resonance stabilization of compound itself.
Electrostatic repulsion between negatively charged phosphate oxygen atoms favor separation of the phosphates.

Question 63

What has the highest energy phosphate bond found in organisms?

Answer 63

PEP (phosphoenolpyruvate) which is involved in ATP synthesis in glycolysis