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Bioc 221 > BIOC 221 - Midterm #2 (advanced editor) > Flashcards

BIOC 221 - Midterm #2 (advanced editor) Flashcards

1
Q

Feedforward Activation ensures that?

A

act in concert to overall goal of E production

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2
Q

An Allosteric Inhibitor does what to an enzyme?

A

binds to enzyme, changes its conformation and changes its substrate affinity (Km)

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3
Q

Bypass 1 - Reciprocal Regulation of Glucose Metabolism

Pyruvate –> ?

Pyruvate Carboxlyase vs PDH complex

A

Acetyl CoA

  • stimulates pyruvate carboxylase (GNG)
  • inhibits PDH complex (CAC)

ATP & NADH

  • inhibits Acetyl-CoA from entering CAC
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4
Q

Reciprocal Regulation is important for two closely parallel pathways because?

direction of reaction is governed by?

A

it prevents concurrent activity which would waste ATP

ΔG (free energy change)

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5
Q

The action of an inhibitor or activator has what effect on:

a) reversible reactions

b) irreversible reactions

A

a) would speeds/slows reverse and forward reaction at same rate (same effect on both)

b) changes overall direction of parallel pathways

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6
Q

Bypass 2 - Reciprocal Regulation of Glucose Metabolism

F6P <–> F16BP

PFK-1 vs FBPase-1

  • inhibited/activated by?
A

PFK- 1

Inhibited by: ATP, citrate

Activated by: ADP, AMP, F26BP

FBPase

inhibited by: AMP, F26BP

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7
Q

**Fructose-2,6-Biphosphate **

Importance? (2)

A

Potent allosteric regulator of PFK-1 and FBPase-1

  • mediator of hormonal regulation of glycolysis and gluconeogenesis
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8
Q

High [F26BP] leads to?

A

Glycolysis increase

PFK-1 - Km decreases

Gluconeogenesis decrease

FBPase-1 - Km increases

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9
Q

How is cellular [F26BP] regulated?

A

Glucagon and Insulin

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10
Q

Effects of

**a) Glucagon **

**b) Insulin **

on blood [glucose]

A

a) raises
b) lowers

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11
Q

F26BP is produced under?

activates? suppresses?

formed by?

inhibited by?

A

normal glucose levels

PFK-1 (glycolysis)

FBPase-1 (gluconeogenesis)

PFK-2

glucagon

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12
Q

F26BP <—> ___

forward and reverse reaction catalyzed by?

A

F26BP -> F6P : FBPase-2

F6P –> F26BP : PFK-2

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13
Q

low blood glucose levels?

A

Pancreas produces Glucagon

Glucagon lowers [F26BP]

low [F26BP] leads to: PFK-1 activation & FBPase-1 inhibition

Glycolysis inhibited

Gluconeogenesis activated

Blood glucose replenished

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14
Q

When glucose is needed?

(4) steps

A

(1) Glucagon
(2) ↓[F26BP], **↑ FBPase-2, ↓PFK-2**
(3) ↓PFK-1, **↑FBPase-1 **
(4) ↑Glycolysis, ↓GNG

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15
Q

When glucose is in excess?

(4) steps

A

↑↓

(1) Insulin
(2) ↑[F26BP], ↓FBPase-2, ↑PFK-2
(3) ↑PFK-1, ↓FBPase-1
(4) ↑Glycolysis, ↓GNG

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16
Q

PFK-2 and FBP-2

A

Bifunctional protein

Glucagon(↑cAMP) - ↑ FBPase-2 (phosphorylated) - ↑GNG

Insulin - ↑ PFK-2 (OH group - dephos) - ↑ Glycolysis

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17
Q

PFK-2 & FBP-2

phosphate group - importance?

A

a phosphate group changes the shape of an enzyme and can alter substrate binding

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18
Q

Cellular Respiration

A

aerobic phase of catabolism where nutrients (sugar, FAs, aa’s) are oxidized to H2O and CO2

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19
Q

CAC - **localization **

A

glycolysis in cytosol

Pyruvate enteres mitochondria to be metabolized further by PDH and CAC

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20
Q

Mitochondrial Compartments

  • Matrix
  • Outer Membrane
  • Inner Membrane Infoldings (Cristae)

`

A

Matrix - PDH complex, enzymes of CAC (also FA ox. and aa metabolism)

Outer Membrane - large channels (leaky)

Inner Membrane Infoldings (Cristae) - contains ETC , major permeability membrane

  • contains transporters
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21
Q

Acetyl-CoA production from ____ by ____

A

Pyruvate

PDH complex

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22
Q

Degradation of 1 glucose to pyruvate via anaerobic glycolysis yields __ ATP.

A

2 ATP

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23
Q

Anaerobic glycolysis only yields 2 ATP.

A much higher yield can be obtained by subsequent?

A

complete oxidative degradation of pyruvate to CO2 and H2O by PDH complex making Acetyl-CoA, then CAC (to CO2) and then ETS

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24
Q

Under aerobic conditions, fate of pyruvate?

A

converted to acetyl-CoA and oxidized to CO2 in CAC

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25
Q

Pyruvate Oxidation to Acetyl-CoA and then CAC

Location?

A

occur in mitochondria

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26
Q

Since glycolysis occurs in cytosol and conversion of pyruvate to ac-CoA and CAC is in mitochondria …

A

pyruvate needs to be transported from cytosol to mitochondrial matrix across two mito. membranes

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27
Q

Inner vs Outer Membrane Transport

A

Inner: highly selective, has specific carrier systems for specific metabolites

Outer: non-specific pores that allows free passage of small metabolites

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28
Q

How does pyruvate get into mitochondria?

A

shuttled into mitochondria by a specific carrier system in exchange for hydroxide ion

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29
Q

Acetyl-CoA

  • importance to metabolic pathways (specifically CAC and glycolysis)
A

initiator of CAC

link between glycolysis and CAC

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30
Q

Pyruvate → Acetyl-CoA

catalyzed by?

cofactors?

A

catalyzed by **Pyruvate Dehydrogenase Complex (E1 + E2 + E3) **

CoA-SH, NAD+, TPP, Lipoate, FAD

CO2 and NADH produced

IRREVERSIBLE

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31
Q

In vertebrates, glucose production from?

A

even numbered FAs impossible

Odd numbers FA’s produce propionyl-CoA (3C) which is converted to pyruvate via succinyl-CoA and OAA

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32
Q

PDH complex

A

multi enzyme complex

series of intermediates remain bound to enzyme molecules

easy flow of intermediated from one active site to another during sequential reactions **(substrate channeling) **

complex, well coordinated regulation

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33
Q

PDH complex - (5) coenzymes

A

lipoamide

Vit B1- thiamine (TPP)

B2 - riboflavin (FAD)

B3 - niacin (NAD)

B5 - pantothenic acid (part of CoA-SH)

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34
Q

Reactions of PDH Complex

(1)

A

pyruvate decarboxylated

remaining hydroxyethyl (2C) group is attached to TPP in E1

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35
Q

Thiamin Pyrophosphate (TPP)

derivative of?

deficiency?

A

derivate of thiamine (vitamin B1)

nutritional deficiency –> Beriberi (loss of neural function)

  • especially affects brain which usually obtains all E from aerobic oxidation of glucose (that includes ox. of pyruvate)
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36
Q

Mechanism of TPP

A

electron sink

H+ dissociates from C between N and S to yield carbanion

e-deficient keto C of pyruvate is attacked by carbanion

then decarboxylation facilitated by e delocalization

2C hydroxyethyl group is now attached to TPP in E1

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37
Q

Reaction (2) of PDH complex

A

Hydroxyethyl (2C) group transferred to lipoamide and is concomitantly oxidized to acetyl group in E1

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38
Q

Swinging Arm - Lipoamide

A

long, flexible arm links lipoamide to E2 (core of the complex) allowing dithiol of lipoamide to swing from one active site to another

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39
Q

Reaction (3) of PDH complex

A

acetyl group is transfered from lipoamide to CoA (in E2)

at the same time, lipoamide is reduced

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40
Q

Reaction (4) of PDH complex

A

dihydrolipoamide is reoxidized to disulfide (-S-S-) form and E3-disulfide is reduced

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41
Q

Reaction (5) of PDH complex

A

the -SH group of E3 are reoxidized by mechanism in which FAD funnels 2e to NAD+ yielding NADH

FAD appears to function as an e conduit (channel)

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42
Q

Summary of PDH complex Reactions

A

Pyruvate decarboxylated & oxidized by NAD+ to acetyl in acetyl-CoA (by now 2C of glucose are lost as CO2)

Free E released during pyruvate ox. is partially stored in NADH & thioster bond in acetyl-CoA

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43
Q

Acetyl-CoA - central to metabolism because?

A

can easily donate acetate based on its high E thioester linkage

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44
Q

The CAC, what for?

A

Continuation of glucose oxidation to CO2

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45
Q

From 1 glucose to acetyl-CoA, we have obtained __ ATP and __ NADH during glycolysis and what from PDH rxn?

A

2 ATP and 2 NADH from glycolysis

2 NADH from PDH reaction

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46
Q

Why is acetyl-CoA the central hub of energy metabolism?

A

degredation of all nutrients (carbs, many aa’s and fat) comes to acetyl-CoA

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47
Q

The basic idea of the CAC`

A

releasing remaining 2 carbons (originally from glucose) in acetyl-coA as CO2 and retaining the free E in the form of ATP, NADH, FADH2

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48
Q

Chemistry of CAC

A

the 2 C in ac-CoA arent directly converted to CO2 (chemically unfeasable).

As the wheel turns, we lose 2CO2 through ox. & decarboxylation per 1 ac-CoA that enters the wheel

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49
Q

CAC - rxn 1

A

Acetyl-CoA + OAA –> citrate

In: H2O Out: CoA-SH

catalyzed by: citrate synthase

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50
Q

CAC - rxn 1

reaction mechanism

A

Citrate synthase: aldol & hydrolysis

binding of OAA to citrate synthase causes a conformation change that opens ac-CoA binding site (induced fit)

transient intermediate: citroyl-coA

citrate is a tricarboxylic acid

-∆G endergonic b/c its irreversible rxn - regulation point

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51
Q

Citrate Synthase Regulation

A

inhibited by:

  • high ATP/ADP and NADH/NAD+ ratios

(high ATP and NADH indicate high E supply for cell)

  • succinyl-CoA (feedback inhibition)
  • citrate (product inhibition)
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52
Q

CAC - rxn 2

A

reversible hydration

Citrate –> [cis-aconitate] –> Isocitrate

H2O out then H2O in

catalyzed by: Aconitase (aconitate hydratase)

2˚ alcohol to 3˚ alcohol

(isomerization)

ENDERGONIC (+ΔG)

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53
Q

CAC - rxn 2

mechanism

A

reversible hydration

3˚ alcohol to 2˚ alcohol

+∆G: Isocitrate is quickly consumed in cell (mass action)

contains Fe-S cluster that aids rxn & binds substrate

intermediate enol compound - cis-Aconitate

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54
Q

CAC - rxn 2 : chemical logic

A

Citrate has 3 –COO– groups, which are almost fully oxidized and ready to be removed as CO2.

easiest way to lose CO2 is through ß-keto decarbox

citrate has no keto , just 1 OH

OH needs to be oxidized to keto

OH group of 3˚ alcohol cant be converted to keto so must convert to a 2˚

this step sets up for oxidation and (facile) ß-keto decarbox in following steps

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55
Q

CAC - rxn 3

A

isocitrate –> α-ketoglutarate

NAD(P)+ -> NAD(P)H + H+

Isocitrate dehydrogenase

1st oxidative decarbox. (β-keto)

enol intermediate tautomerized to α-ketoglutarate

exergonic

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56
Q

CAC- rxn 3

mechanism

A

oxidation of C2 alcohol of isocitrate w/ reduction of NAD+ to NADH

  • followed by β-keto decarbox. of central carboxyl
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57
Q

Reaction 3 of CAC is identical to which other reaction?

A

6-phosphogluconate DH rxn in oxidative phase of PPP

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58
Q

α-KG is an important metabolite in?

A

amino acid metabolism

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59
Q

CAC - rxn 4

A

α-KG –> Succinyl Co-A

α-KG dehydrogenase

CoA-SH, NAD+

2nd oxidative decarbox.

exergonic

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60
Q

CAC - rxn 4 - mechanism

A

α-keto decarbox.

uses same enzymes as PDH and cofactors TP, lipoate, FAD

E released from ox. & decarb. conserved in NADH and succinyl-CoA thioester bond

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61
Q

By the 4rth step of the CAC, we’ve lost…

the remaining steps are to?

A

2 CO2

to regenerate OA to complete cycle

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62
Q

CAC- rxn 5

A

Succinyl CoA–> Succinate

GDP + Pi -> GTP + CoA-SH

Succinyl-CoA Synthetase

exergonic

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63
Q

CAC - rxn 5 : mechanism

A

synthesis of GTP

thioester cleaved driving substrate-level phosphorylation

exergonic (barely)

Pi acts as Nu allowing CoA-S to leave

Pi eventually passed to GDP to form GTP

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64
Q

CAC- rxn 6

A

Succinate –> Fumarate

Succinate DH (SDH)

prosthetic group: FAD (bound to enzyme)

ΔG ~ 0 kJ/mol

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65
Q

CAC - rxn 6 mechanism

A

enzyme-linked FAD is e acceptor (better acceptor than NAD+)

E-FADH2 is reoxidized by coenzyme-Q in ETC

this is why it is the only membrane bound enzyme in CAC

(embedded in mito. inner membrane allowing it to be part of complex II of ETC)

FeS clusters provide direct pathway for e’s to ETC leading to synthesis of approx. 1.5 ATP

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66
Q

CAC - rxn 7

A

Fumarate –> L-malate

anti-hydration (add H2O) - OH and H on opposite side

fumurate hydratase

stereospecific (only produces trans L-malate)

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67
Q

CAC - rxn 8

A

oxidation of malate (redox)

L-malate –> Oxaloacetate (regenerated!)

endergonic

Malate DH

(reduction of NAD+)

reverse rxn in gluconeogenesis (OA malate shuttle)

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68
Q

If Rxn 8 of CAC is endergonic, how does it occur?

A

driven by mass action (product depletion)

  • OAA taken up quickly in cycle by highly exergonic citrate synthase rxn

[OAA] < 10-6 M makes rxn favorable

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69
Q

Which Rxns in CAC are irreversible?

A

rxn 1) Ac-CoA –> Citrate
rxn 3) Isocitrate –> α-KG

rxn 4) α-KG –> Succinyl-CoA

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70
Q

Prochiral

A

molecules that can be converted from achiral to chiral in a single step

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71
Q

Glucose C1 & C6 become:

A

CH3 of pyruvate & ac-CoA

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72
Q

Glucose C2 & C5 become:

A

Carbonyl (C=O) C of pyruvate & ac-CoA

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73
Q

Glucose C3 & C4 become:

A

lost as CO2 during PDH

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74
Q

If Carbonyl C is radiolabelled, at what step does the radioactivity split?

Why?

A

Conversion of Succinyl-CoA to Succinate

because Succinate symmetrical and the 2 COO- groups (C1 & C4) are chemically equivalent

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75
Q

When are radiolabelled carbonyl C lost as CO2 in CAC?`

A

in the second round

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76
Q

When is methyl (CH3) C of ac-CoA lost as CO2 in CAC?

A

methyl C survices 2 complete cycles but 1/2 of whats eft exits cycle on each turn after that

2CO2 that are released during 3rd round are radiolabeled

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77
Q

+E° ?

A

accepts e’s - gets reduced

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78
Q

-E° ?

A

gives e’s - gets oxidized

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79
Q

Products of 1 turn of CAC

A

3 NADH

1 ATP

1 FADH2

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80
Q

The Amphibolic nature of the CITRIC ACID CYCLE

A

metabolic pathway involved in both anabolism and catabolism

  • much of CAC evolved before aerobes
  • used for anabolism in anaerobes
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81
Q

The CAC intermediates usually remain constant as a result of?

A

Anaplerotic reactions that replenish CAC intermediates `

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82
Q

Anaplerotic reactions of the CAC (3)

A

1) pyruvate + HCO3- + ATP <=pyruvate carboxylase=> OAA + ADP + Pi

2) PEP + CO2 + GDP <=PEP carboxylase=> OAA + GTP
3) pyruvate + HCO3- + NAD(P)H <=malic enzyme=> malate + NAD(P)+

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83
Q

To keep the cell in stable steady state & to avoide wasteful overproduction, the Citric Acid Cycle is regulated by (3) ?

A

1) substrate availability
2) product inhibition & allosteric feedback inhibition
3) covalent modification

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84
Q

(4) points of CAC regulation

A

1) PDH complex (pyruvate –> ac-CoA)
2) citrate synthase (ac-CoA + OAA –> Citrate)
3) Isocitrate DH (Isocitrate –> a-KG)
4) a-KG DH complex (a-KG –> Succinyl-CoA)

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85
Q

CAC regulation by: Substrate Availability

A

substrate availability varies w/ cell metabolic state and [ac-CoA] & [OAA] controls citrate synthase

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86
Q

CAC regulation by product inhibition

A

a) high [NADH]/[NAD] can inhibit all Dehydrogenases by mass action & NADH competes with NAD+ for binding
b) PDH complex - ac-CoA competes with CoA for binding to E2

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87
Q

CAC regulation by allosteric feedback inhibition

A

b) Citrate Synthase & a-KG DH
- by NADH and/or ATP

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88
Q

CAC regulation by covalent modification (Ca2+ signals)

A

PDH, Isocitrate DH, a-KG DH - regulated by calcium

release of Ca2+ stored in sarcoplasmic reticulum induced by neurons (activated by Ca2+)

  • contraction signal
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89
Q

CAC regulation by covalent modification

A

phosphorylation of PDH E1 by pyruvate DH kinases (PDK’s) inactivates enzyme

1) PDK’s activated by ATP (signalling excess E)

OR

2) during low [glucose], glucose required by brain so catabolism blocked in muscle mito by increase PDK activity that phosphorylates & shuts down PDH

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90
Q

If there is alot of citrate in mitochondria, it can be transported to cytosol, causing?

A

signals for FA synthesis

inhibition of: PFK-1

converted to ac-CoA & OA by citrate lyase

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91
Q

What happens when cells energetic needs are met?

A

(high [ac-CoA/citrate/ATP] favors glucose & glycogen syntheses

inhibition of CAC - accumulates ac-CoA –> FA synthesis

excess ATP inhibits Ox. phosphorylation, NADH accumulates

excess pyruvate is converted to glucose (GNG) –> glycogen synthesis

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92
Q

(2) hormones signal when metabolic E is required

A

1) Glucagon - low glucose levels
2) Epinephrine - need immediate energy

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93
Q

When either glucagon or epinephrine are secreted..

A

adenyl cyclase is activated, triggering cascade response

  • cAMP acts as second messenger
  • activated PKA phosphorylates lipase & perilipin

perilipin-P allows lipse-P access to lipid droplet surface

  • lipase-P converts TAG’s to FA’s

transported by serum albumin to skeletal muscle, heart, kidney

enter cells by transporter

ß-oxidation to CO2 yielding ATP

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94
Q

the (2) degredation products of TAGs

A

1) free fatty acids
2) glycerol

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95
Q

Fate of degradative product of TAGs: Fatty acids

A

ß-oxidation in mitochondria (in animals)

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96
Q

small FAs can diffuse freely across mitochondrial membrane. How do larger FAs enter mitochondria?

A

Carnitine shuttle

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97
Q

Carnitine shuttle - (1)

A

activation by acyl-CoA synthetases at OMM

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98
Q

Carnitine shuttle (1) - activation by acyl-CoA synthetases at OMM

A

leaving group activation

carboxylate ion is adenylated by ATP and PPi is hydrolyzed to 2 Pi

CoA-SH thiol attacks, AMP leaves

forming fatty acyl-CoA

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99
Q

Carnitine Shuttle - (2)

A

transfer of acyl-CoA to matrix

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100
Q

Carnitine shuttle (2) - transfer of acyl-CoA to matrix

A

fatty acyl group transferred to carnitine by carnitine acyl-transferase I

transport : IMS –> matrix through acyl-carnitine transporter

fatty acyl transfer from carnitine back to CoA to regenerate fatty acyl-CoA in matrix

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101
Q

Why isnt fatty acyl-CoA just transported into matrix through a certain transporter?

A

to keep 2 seperate pools of CoA and fatty acyl-CoA (1 in mitochondria, 1 in cytosol)

  • have different functions
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102
Q

Functions of

1) cytosolic CoA
2) mitochondrial CoA

A

1) biosynthetic (membrane lipids)
2) catabolic (ox. degredation of pyruvate by PDH, FAs, AAs)

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103
Q

Malonyl-CoA inhibits?

A

carnitine acyltransferase I

malonyl-CoA is 1st intermediate for FA synthesis from acetyl-CoA

  • high [malonyl-CoA] indicates time for FA synthesis & inhibits entry of FAs into mitochondria
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104
Q

Fate of degredative product of TAG: glycerol

A

adipocytes lack glycerol kinase

glycerol shuttled to liver via blood & converted to G3P & DHAP for glycolysis or GNG

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105
Q

(3) stages of Fatty Acid Oxidation

A

1) oxidative conversion of 2C units into ac-CoA w/ concomitant generation of NADH
2) oxidation of ac-CoA into CO2 via CAC w/ concomitant generation of NAD+ & FADH2
3) generates ATP from NADH & FADH2 via ETC

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106
Q

Stage (1) - ß-oxidation

A

every other C is converted to C=O

allows Nu attack of CoA-SH

each round produces: 1 NADH, 1FADH2, 1 ac-CoA (2 in last round)

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107
Q

ß-Oxidation - step 1

A

dehydrogenation of alkane to alkene by acyl-CoA DH (AD) on the IMM

FAD = cofactor as e acceptor

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108
Q

ß-oxidation - step 2

A

hydration of alkene by enoyl-CoA hydratase

H2O added across double bond yields alcohol

stereospecific - only L

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109
Q

ß-oxidation - step 3

A

dehydrogenation of alcohol by ß-hydroxyacyl-CoA DH

NAD = cofactor as hydride acceptor

only L-isomers of hydroxyacyl CoA

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110
Q

ß-oxidation - step 4

A

transfer of FA chain
by acyl-CoA acetyltransferase

carbonyl C in ß-ketoacyl-CoA is electrophilic

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111
Q

Which 3 successive enzymes in either a pathway or cycle are analagous to 3 enzymes in ß-oxidation and why?

A

succinate DH

fumarase

malate DH

(oxidation of ß CH2 to alcohol then carbonyl C=O )

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112
Q

FA synthesis takes place in?

A

cytosol

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113
Q

FA degredation takes place in?

A

mitochondrial matrix

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114
Q

FA synthesis

A

FA chain elongated by 2C (acetate) units

activated donor of 2C units is (3C) malonyl-ACP

intermediates are attached to acyl carrier protein (ACP)

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115
Q

1st committed step in FA synthesis

A

formation of malonyl-CoA from ac-CoA & HCO3-

(1 ATP used)

acetyl-CoA + HCO3- –> malonyl-CoA

catalyzed by acetyl-CoA carboxylase (ACC)

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116
Q

FA synthesis is similar to reverse of FA degredation except (2)?

A

1) NADPH is used
2) stereochemistry of hydroxylated intermediate is reverse

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117
Q

FA synthesis: supply of acetyl-CoA

A

acetyl-CoA is synthesized in matric

IMM is impermeable to acetyl-CoA so acetyl-CoA units are shuttled out of matrix as citrate

shuttle also substitutes a NADPH for an NADH which is needed for synthesis

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118
Q

Regulation of Fatty Acid Oxidation

A
  • *compartmentalization -**
  • synthesis of TAGs* - cytosol, liver, adipocytes, intestine

oxidation to acetyl-CoA - mitochondria

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119
Q

rate of ß-oxidation is controlled by?

A

rate at which acetyl-CoA is transported into mitochondria by carnitine acyltransferase I

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120
Q

Regulation of FA biosynthesis (3)

A

1) allosteric regulation of ACC
2) regulation of gene expression by FAs
3) hormonal regulation of enzymes by covalent mod.

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121
Q

regulation of FA synthesis

1) allosteric regulation of acetyl-CoA carboxylase

A

citrate is positive effector (feedforward activation)

palmitoyl-CoA is negative effector (feedback inhibition)

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122
Q

Regulation of FA synthesis

3) hormonal regulation of enzymes by covalent modification (ACC)

A

Acetyl-CoA carboxylase

high blood glucose: insulin: activates Pase, dephosphorylates & activates ACC, malonyl-CoA inhibits ß-oxidation

low blood glucose: glucagon: activate kinase, phosphorylatres & inactivates ACC, malonyl-CoA not made, ß-oxidation to produce ATP , acetyl-CoA to CAC to make more ATP

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123
Q

(2) enzymes that are key to coordination of FA metabolism

A

1) carnititine acyltransferase 1 & acetyl-CoA carboxylase

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124
Q

Citrate is an effector for…

A

PFK-1 : inactivates

ACC: activates

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125
Q

How does citrate regulate?

A

citrate shuttle

xs mitochondrial ATP & acetyl-CoA increases transport of citrate out of mitochondria to cytosol

citrate turns down glycolysis in cytosol and switches on FA biosynthesis (increases ACC)

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126
Q

Malonyl-CoA as an effector

A

malonyl-CoA shuts down ß-oxidation

1st intermediate in FA synthesis

shuts down transport step (inhibits carnitine acyltransferase I)

good example of compartmentalization

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127
Q

During fasting or carb starvation…

what is depleted in liver?

rates of pathways?

A

OAA

glycolysis rate is low so supply of precursors for replenishing OAA is cut off and OAA is siphoned off into GNG to maintain blood glucose

128
Q

During fasting or starvation… the lack of OAA (being used for GNG) impedes entry of acetyl-coA.

What happens with Acetyl-CoA?

A

Acetyl CoA accumulates in liver mitochondria is converted to ketone bodies: acetoacetate, B-hydroxybutryate & acetone? which are released into bloodstream for organs other than liver (heart, brain) to use as fuels

129
Q

Ketone Body Production - rxn 1

A

2 Acetyl-CoA condense to form **acetoacetyl-CoA **(catalyzed by thiolase) - reverse reaction in B-oxdn

(coA-SH leaves)

130
Q

Ketone Body Production

2 Acetyl-CoA –> Acetoacetyl-CoA –> ? –> ?

A

addition of another ac-CoA forms HMG-CoA

cleaved to form acetoacetate & acetyl-CoA

131
Q

Ketone Body Production

2 Acetyl-CoA –> Acetoacetyl-CoA –> HMG-CoA –> acetoacetate + acetyl-CoA –> ?

2 fates of acetoacetate

A

acetoacetate **reduced **to B-hydroxybutyrate

or

spontaneously decarboxylated to acetone (exhaled)

132
Q

Ketone bodies

properties

A

water soluble

can be transported in blood to other tissues including brain

enters CAC or used to make myelin

133
Q

Ketone bodies transported to extraheptatic tissues..then?

A

converted back to acetyl-CoA

B-hydroxybutyrate can produce 1 NADH, 2 ac-CoA in CAC & ETC

134
Q

Why can’t liver use ketone bodies for fuel?

A

liver doesnt have B-kat that converts B-hydroxybutryate to acetoacetyl-CoA (then converted to 2 ac-CoA by thiolase)

135
Q

What uses acetoacetate in preference to glucose?

A

heart muscle & renal cortex

136
Q

If acetoacetate is overproduced?

A

B-HB will lower pH (ketoacidosis) of blood

137
Q

Why go through HMG-CoA?

A

to prevent facile B-decarbox. during transport

138
Q

why not ship out ac-CoA rather than b-hydroxybutyrate

A

liver has only limited supply of CoA which is needed for B-oxdn

139
Q

(3) reasons why AAs need to be ox. degraded

A

1) protein turnover
2) high protein diet
3) starvation or untreated diabetes

140
Q

Digestion of Protein

(2) steps

A

protein ingestion stimulates gastrin (hormone) secretion which stimulates release of pepsinogen & **HCl **

141
Q

Release of HCl & Pepsinogen stimulated by Gastrin

A

HCl - denatures proteins (antiseptic)

Pepsinogen - precursor of pepsin that cuts proteins into peptide fragments & AAs

  • causes cholecystokinin secretion in duodenum
142
Q

Pepsinogen causes cholecystokinin secretion in duodenum which …

A

stimulates release of zymogens to pancreas

other proteases are released from pancreas ** **

143
Q

low pH in intestine triggers..

A

**secretin **release in blood, stimulating HCO3- (bicarbonate) release to neutralize pH

144
Q

Pepsinogen is activated by?

A

autoproteolytic cleavave at lower pH

(this stimulates secretin to stimulate release of HCO3 to neutralize pH)

145
Q

why synthesize digestive enzymes as inactive zymogens?

A

to protect **exocrine cells **from **proteolytic attack **

146
Q

Chymotrypsinogen & Trypsinogen (inactive)

A

proteases (neutral pH optimum) that are released from pancrease by action of **cholecystokinin **and work in small intestine at neutral pH

  • activated by proteolytic cleavage
147
Q

Trypsin

inhibited in pancreas by?

A

Pancreatic trypsin inhibitor

148
Q

carboxy & aminopeptidases

A

participate in degrading shorter peptides

carboxy and amino terminal ends are removed one at a time

free amino acids are transported through intestinal mucosa through blood and to liver

149
Q

once the amino acids from ingested protein are transported to liver…

A

process 1) transamination

150
Q

Transamination - process 1

A

AA is converted to a-keto acid and a-KG accepts amino group to become glutamate

(cytosol in mammals)

151
Q

After transamination (aa becomes a-keto acid and a-KG accepts amino group to become glutamate)?

A

a-keto acid formed can go to CAC (pyruvate, OAA)

glutamate transported to liver mitochondria for deamination

152
Q

Process (2)

A

**deamination **of glutamate to a-KG in liver mitochondria

153
Q

Process (3)

A

ammonia from deamination of glutamate transported as amide N of glutamine

154
Q

IN muscle.. xs ammonia?

A

transferred to pyruvate –> alanine

155
Q

(3) process of amino group transfer and transport

A

1) transamination
2) deamination
- glutamate dh (ox. deam.)
- glutaminase (hydrolyt. deamn)
3) glutamine synthetase

156
Q

Process 1) transamination reactions - aminotransferase

purpose?

A

to collect aa groups as l-glutamate from many different aas

glutamate functions as amino group donor for biosynthetic pathways or excretion (urea cycle)

157
Q

Transamination reaction

2) steps

PLP acts as?

A

bimolecular pingpong

1) amino acid binds, donates amino group to **PLP **& leaves as a-keto acid
2) another a-keto acid binds & accepts amino group from **PLP, **leaves as amino acid
* PLP acts as intermediate amino group carrier*

158
Q

Deamination reaction - enzyme?

process?

A

glutamate DH

glutamate from transamination rxn is transported to mitochondria for ox. deamin. to give a-KG & NH4+

159
Q

Glutamate DH - importance?

A

only enzyme that can use NAD and NADP

at important intersection of C & N metabolism

regulated by array of allosteric effectors

160
Q

Glutamate DH

allosteric effectors

A

inhibitors: GTP
activators: ADP

161
Q

Amino group transport

A

glutamine synthetase

ammonia requires conversion before transport from extrahepatic tissues to blood

free ammonia + glutamate –> glutamine

162
Q

uses of glutamine

A

1) purine synthesis
2) xs is transported to **liver/kidney **& deaminated by **glutaminase **(in liver mito)

163
Q

Glucose-Alanine Cycle

A

ala can carry NH4+ & carbon skeletons of pyruvate between muscles & liver

164
Q

Glucose-Ala Cycle

  • details
A

muscle protein is broken down to AA to be used for fuel

muscle **aminotranferase **uses pyruvate (from glycolysis) as amino acceptor to make alanine

**Alanine **travels to liver - transamination -> pyruvate -> GNG -> glucose -> back to muscle

amino group from transam back to pyruvate goes to urea cycle

165
Q

**Cori Cycle **

A

moves pyruvate, lactate, ammonia to liver

166
Q

NH4 formed in other tissues reaches the liver how?

A

transported to liver as amide of glutamine

167
Q

Cori Cycle & Glu-Ala Cycle are ___ pathways through which ___ and ___ exchange ___ ___

A

multiorgan

liver and muscle

metabolic intermediates

168
Q

How is amino group (or ammonia) used or eliminated?

1) aquatic species
2) plants
3) reptiles/birds
4) synthesis
5) conversion

A

1) secrete
2) recycle
3) excrete uric acid in eggs (solid b/c it precipitates)
4) synthesis of amino acids
5) converted to urea

169
Q

**Structure of Urea **

  • each part derived from?
A

H2N-(C=O)-NH2

1 NH2 derived from deamin. of glutamine or glutamate in mito through carbamoyl phosphate

other NH2 from aspartate

central carbon from bicarbonate also through carbamoyl phosphate

170
Q

Urea Cycle - Diagram

A

look at diagram

171
Q

Carbamoyl-P amino group can come from the ammonia that has been transported to liver by? (4)

A

1) ammonia - portal vein/bac ox. of aa
2) glutamine - extrahepatic
3) aa’s - (glutamate)
3) alanine (muscle)

172
Q

Synthesis of Carbamoyl-P

A

ATP + Bicarbonate (HCO3-): **LG activation **

bicarbonate is phosphorylated

ammonia displaces P group to make carbamate

carbamate phosphorylated to yield carbamoyl-P

173
Q

Activation of carbamoyl-P requires ….

174
Q

Once carbamoyl-P is made…

A

enters in urea cycle with ornithine to make citrulline

ornithine + carbamoyl-P –> citrulline + Pi

175
Q

after carbamoyl-P joins with ornithine to make citrulline

A

citrulline is transported out of matrix to cytosol

176
Q

after citrulline is transported from matrix to cytosol

A

**arginosuccinate synthetase **

carbonyl of citrulline attacks AMP of ATP - LG=PPi

addition of aspartate to citrullyl-AMP - LG=AMP

activation of ureido oxygen of citrulline sets up addition of aspartate to form arginosuccinate

177
Q

Aspartate-arginosuccinate shunt

A

link b/w urea cycle & CAC

178
Q

Aspartate-Arginosuccinate Shunt - link b/w Urea Cycle & CAC

A

In urea cycle: OAA -> asp -> fumarate (in cytosol) -goes into mitochondria to form 1 NADH when converted to OAA

179
Q

UREA CYCLE - overall

A

NH4+ + HCO3- + aspartate + 3ATP –> urea + fumarate + 2ADP + AMP + 4Pi

180
Q

For Urea Cycle - 3 ATP used where?

A

2 for carbamoyl-P and 1 for citrullyl-AMP

181
Q

How does the pathway interconnections between CAC & Urea cycle reduce energetic cost of Urea Cycle?

A

**Aspartate **

**- **needed for cytosolic conversion of citrulline to **arginosuccinate **

  • produced when **OAA **accepts amino group from glutamate

**fumarate **to **OAA **produces 1 NADH

*Glutamate DH rxn *also produces 1 NAD(P)H (glu–> a-KG)

(1 NADH = 2.5 ATP)

182
Q

All amino acids become ..? (2)

A

CAC intermediates

Ac-CoA

183
Q

CAC intermediates are (3)

A

1) diverted to GNG (forming glucose)
2) diverted to ketogenesis (formation of ketone bodies)
3) completely oxidized to CO2 & H2O

184
Q

Genetic disorders related to AA metabolism

A

most cases of genetic defects in aa metabolism lead to defective neural development & mental retardation

  • most aa’s are neurotransmitters, precursors, antagonists
185
Q

Phenylketonuria

A

Phe hydroxylase mutation

Phe may compete w/ other amino acids for transport across blood brain barrier

186
Q

Alternative pathways for catabolism of Phe in PKU

(when there is Phe buildup)

A

Phe + pyruvate –> Phenylpyruvate + alanine (aminotransferase)

phenylpyruvate –> phenylacetate + phenyllactate

all 3 products build up in tissues, blood & urine

187
Q

Treatment for PKU

A

limiting Phe intake to levels barely adequate to support growth

Tyrosine is an essential nutrient for individuals with PKU must be supplied in their diet

188
Q

Location of

1) glycolysis
2) PDH rxn
3) CAC
4) GNG
5) FA oxdn
6) FA synthesis

A

1) cytosol
2) cytosol
3) mito. matrix
4) cytosol (except one rxn in lumen of ER)
5) mitochondria
6) cytosol

189
Q

Chemiosmotic Theory

A

ATP synthesis & electron transport are coupled by H+ gradient across mito membrane

190
Q

Overview of ETC

A

1) flow of e’s through **membrane-bound carriers **
2) exergonic e flow couples to endergonic H+ transport against [c] gradient
3) H+ transport down [c] gradient through specific protein channels provides E for ATP synthesis
4) ATP synthase couples H+ flow to ADP phosphorylation

191
Q

Other Electront Carrying Molecules that transfer e’s through membrane

(3)

A

1) Ubiquinone (coenzyme Q)
2) cytochrome
3) iron-sulfur proteins

192
Q

Ubiquinone (coenzyme Q)

A

hydrophobic, lipid-soluble

benzoquinone + isoprenoid side chain

  • shuttles e through the membrane (lateral diffusion)

carries both e- & H+

193
Q

Cytochromes

A

proteins with iron-containing heme prosthetic groups

reduction potential depends on heme environment (aa’s surrounded - electrostatic effects)

194
Q

Hemes a & b vs c Heme.

A

a & b: loosely associated with enzyme

c: covalently linked (prosthetic) coenzyme

195
Q

cytochromes a, b & many c are what kind of proteins?

cyt c enzymes are what kind of membranes?

A

integral membrane proteins

peripheral membrane proteins associated through electrostatic interactions w/ the IMM outer surface (on side of IMS)

196
Q

Iron-sulfur Proteins

A

contain iron-sulfur clusters (1 e transfer)

at least 8 in mito ETC

reduction potential

197
Q

Components of ETC

A

Complex :

I - NADH DH

II - Succinate DH (in CAC)

III - Ubiquinone: cyt c oxidoreductase

cytochrome c1

IV - cytochrome oxidase

198
Q

Path of electrons from

1) NADH
2) succinate
3) fatty acyl-coA
4) G3P

A

1) complex 1 : FMN -> Fe-S –> Q
2) Complex II- Succinate oxidized Fumarate: FAD –> Fe-S –> Q
3) Fatty acyl-CoA -> Enoyl CoA : FAD-> FAD-> FAD, Fe-S
4) cytosolic G3P to G3PDH

199
Q

Complex I: A Proton Pump

A

**NADH DH **

NADH Ubiquinone oxidoreductase

transfers e’s from NADH to ubiquinone

coupled rxns:
a) **exergonic: **NADH + H+N+ Q –> NAD+ +QH2

b) **endergonic: **vectorial translocation of 4H+ (per 2e)

matrix (N side) becomes -ve; IMS (P side) becomes +ve charged

proton gradient

QH2 diffuses laterally to complex II

200
Q

Complex II

A

Succinate to Ubiquinone

(succinate - FAD - Fe-S - ubiquinone (Q–>QH2))

complex includes succinate DH

  • only membrane bound enzyme of CAC

not a proton pump

201
Q

Other mitochondrial DHs

A

other substrates of DHs (i.e. **acyl coA DH **) can pass electrons to ETC through ubiquinone

fatty acyl-CoA –> enoyl CoA (1st step in ß-ox)

202
Q

Structure of Complex II (succinate DH)

binding sites

bound

function of heme b

A

binding sites: succinate, ubiquinone

bound: FAD, FeS clusters, hemes

heme b is not in direct path of e transfer but thought to prevent leakage of e’s & conversion of H2O2 to oxygen radicals that will damage tissue

203
Q

Complex III

A

cyt bc1 complex - transfers e’s from ubiquinol (QH2) to cyt c

a) **exergonic: **QH2 + cytc(ox) –> Q + cytc(red)
b) **endergonic: **translocation of 4H+/2e

204
Q

cytochrome c

A

cyt c (soluble) heme accepts e’s from complex III and moves to complex III

205
Q

Cavern of Complex III

A

space inside complex in which Q is free to move from N side of membrane to IMS as it shuttles e- & H+ across IMM

206
Q

How do we transfer 2e’s to a 1e carrier?

(QH2 to cyt c)

(Qcycle)

A

QH2 donates 1 e- to cyt c1 (via rieske)and the **other to Q **(via cyt. b)

2H+ are pumped in this 1st half of the Q cycle

semiquinone radical formed

another QH2 donates 1e to another **cyt c1 **and the other to **semiquinone (Q radical) **(also 2H+ to form QH2)

this pumps another 2H+ to IMS (Pside)

207
Q

Overall rxn of Q cycle

A

QH2 + 2cytc1 (oxidized) + 2H+N ==> Q + 2cytc1 (**reduced) **+ 4H+P

208
Q

The **Q cycle **in 2 stages

FIRST STAGE & SECOND STAGE** **

explain what happens on P & N side

A

first, Q on N side is **reduced **to semiquinone radical

in *second stage, *semiquinone radical is further **reduced **to QH2

on P side: 2 molecules of QH2are oxidized to Q releasing 2H+ per Q (4 overall) into IMS (Pside)

Each QH2donates: 1e to cyt c1 (via rieske Fe-S center)

1e to Q near N side (via cyt b) - uses 2H+ per Q taken from matrix

209
Q

Complex IV

aka?

reactions?

overall?

A

cytochrome oxidase since it transfers e’s from cyt c to oxygen

**exergonic: **4cytred + 4H+N + O2=> 4cyt(ox) + 2H2O

**endergonic: **translocation of 1H+ per 1e

(4H+N –> 4H+P)

overall: 4cytred+ 8H+N + O2 => 4cyt(ox) + 4H+P+ 2H2O

**copper **& **heme **bound proteins are involved in e transport

210
Q

summary of flow of e’s & protons through 4 complexes of respiratory chain

A

e’s reach **Q **through **complexes I & II **

reduced Q (QH2) serves as mobile carrier of e’s & protons

  • passes e’s to **complex III **which passes them to cyt c (mobile link)

**complex IV **transfers e’s from **reduced cyt c **to O2

211
Q

Which complexes have proton flow?

A

e flow through **complexes I, III & IV **are accompanied by proton flow from **matrix **to **IMS **

212
Q

proton pumped in ETC (each complex & overall)

A

**Complex I: **4H+

**Complex III: **4H+

**Complex IV: **2H+

from matrix (N) to **IMS (P) **

OVERALL: 10H+P

213
Q

Overall Rxn of ETC

A

NADH + 11H+N+ 1/2O2 → NAD+ + 10H+P + H2O

214
Q

What is the Chemiosmotic Hypothesis?

A

ATP synthesis & electron transport are coupled by electrochemical gradient across mito. membrane

215
Q

Chemiosmotic Theory - respiration

coupled reactions

what is created?

A

spontaneous (exergonic) e transfer through complexes I, III & IV is coupled to non-spontaneous (endergonic) H+ pumping from matrix

H+ pumping creates electrochemical gradient, **proton-motive force

a **membrane potential **(-ve in matrix) & pH gradient (alkaline in matrix)

216
Q

Electrochemical gradient

A

proton motive force

consists of both: **membrane potential **& **pH gradient **

217
Q

Chemiosmotic Theory - F1FoATP synthase

coupled rxns?

driving force?

A

non-spon. ATP synthesis coupled to spont. H+ transport into matrix

pH & electrical gradients created by respiration = driving force for H+ uptake

H+ returns to matrix via Fo uses up pH & electrical gradients

218
Q

Energy needed to transport solute against conc gradient

A

∆G = RT ln (C2/C1)

C1 < C2 , ∆G > 0

219
Q

Net movement of an electrically **neutral **solute is towards?

A

side of **lower solute **concentration until eq. is achieved

220
Q

Energetics of **ION **transport across membranes

movement of ion without counterion…

A

movement of ion without a counterion results in **endergonic **seperation of +ve and -ve charges, producing electrical potential

221
Q

Energy cost of moving an ion depends on?

A

**electrochemical potential: **the sum of chemical & electrical gradients

∆Gt= RT ln (C2/C1) + ZF∆ψ

222
Q

Direction of net movement of an electrically charged solute is dictated by…

A

a combination of **chemical conc. difference (C2/C1) **

and the **electrical potential (Vm) **across the membrane

net ion movement continues until electrochemical potential = 0

223
Q

Proton Motive Force (PMF)

A

energy stored as proton gradient

protons can flow spontaneously down **electrochemical gradient, **and energy is available for work (ADP –> ATP)

224
Q

Chemiosmosis

A

movement of ions across selectively permeable membrane, down electrochemical gradient

225
Q

Chemiosmotic Model

A

**oxidation & phosphorylation **become obligately coupled (absence of one inhibits the other)

226
Q

How do H+ reenter matrix?

A

IMM is impermeable to H+

H+ can only reenter the matrix through **proton-specific channels (Fo) **

227
Q

What provides E for ATP synthesis?

ATP synthesis catalyzed by?

A

**proton-motive force **that drives H+ back into matrix

F1 complex associated with Fo

228
Q

What happens if **complex I, III, IV or ubiquinone **is blocked?

A

no electron transport

no proton gradient produced

shuts down ATP synthesis

229
Q

What happens when ATP Synthase is blocked? (adding oligomycin)

(4)

A

no ATP produced

no release of proton gradient & PMF builds up

since high [H+] build up is not dissipated, free energy released by oxidn of substrates is not enough to pump any more protons against steep gradient

shut down of electron transport

230
Q

Can we have electron transport without ATP synthesis?

A

uncoupled by uncoupler

231
Q

various inhibitors can be used to demonstrate coupling of (2)

A

1) ETC

and

2) proton gradient with ATP synthesis

232
Q

DNP - what is it?

properties & characteristics

A

a chemical uncoupler of **oxidative phosphorylation **

  • has a dissociable proton & very hydrophobic
233
Q

what does DNP do?

A

carries protons across IMM, dissipating proton gradient

234
Q

What happens to

a) O2 consumption
b) **ATP synthesis **

when succinate is added (or any oxidizable substrate)

A

a) slight increase in slope
b) nothing

235
Q

What happens to

a) O2 consumption
b) ** ATP synthesis **

** **when ADP + Pi are added (after added succinate)

A

a) increase
b) increase

236
Q

What happens to

a) O2 consumption
b) ** ATP synthesis **

when oligomycin is added (after adding succinate, ADP & Pi)

A

a) rate of consumption decreases so slope is same as beginning (slowed)
b) horizontal line (no ATP synthesis)

237
Q

What happens to

a) O2 consumption
b) **ATP synthesis **

when DNP is added (after succinate, ADP, Pi & oligomycin are added)

A

DNP disrupts proton gradient (dissipates)

uncouples

a) increases
b) stays the same (no ATP synthesis - horizontal line)

238
Q

Artificially created PMF for ATP synthesis?

(solution 1 & 2)

A

isolated mito. are first incubated in pH 9 buffer containing **0.1 M KCl **so matrix reaches eq. w/ surroundings (KCl & buffer leak into mito) then resuspended in pH 7 buffer containing **valinomycin **& no KCl

change in buffer creates a different of 2 pH unites across IMM.

outward flow of K+ (carried by valinomycin) down conc. gradient without counterion Cl- creates **charge imbalance across membrane **(matrix = -ve)

sum of **chemical potential by pH difference **& **electrical potential by seperation of charges **is a PMF large enough to support ATP synthesis in absence of oxidizable substrate

239
Q

ATP synthase in humans

A

F-type ATPase

240
Q

F-type ATPase

domains (subunits)

A

large enzyme complex

2 functional domains : F1 & Fo

241
Q

function of F1 domain in F-type ATPase

A

catalyzes ATP synthesis from ADP + Pi

242
Q

Function of **Fo domain **of F-type ATPase

what does ‘o’ of **Fo **stand for?

A

allows H+ to passively diffuse from P to N side (proton pore)

o = oligomycin sensitive

243
Q

When dissociated… F1 is ___, Fo becomes a ____ __

A

F1 = ATPase

Fo becomes H+ pore

244
Q

Mechanism of F2

(experiment)

A

reversiblity & enzyme stabilization

18O exchange experiment showed that when F1 is incubated with ATP & H218O , the Pi contained 3-4 of the 18O, indicating that both ATP hydrolysis & synthesis have occured several times during

reaction is reversible & exchange doesnt require input of energy

245
Q

Keq for ATP hydrolysis

a) in solution
b) on F1

why the huge difference?

A

a) 105
b) 2.4

ATP synthase stabilizes ATP relative to ADP + Pi by binding more ATP more tightly, releasing enough E to counterbalance cost of making ATP

246
Q

ATP can only be released from F1 through..

driven by?

requires?

A

conformational change of F1

proton gradient

requires Fo

247
Q

Major energy barrier in ATP synthesis catalyzed by ATP synthesis

A

release of ATP from enzyme (not formation of ATP)

248
Q

Free-E change for formation of ATP from ADP & Pi in solution is large & (+) but on the enzyme surface…

A

tight binding of ATP provides sufficient binding energy to bring the free energy of E-bound ATP close to that of ATP & Pi so reaction is readily reversibleb

eq constant near zero

249
Q

Where does the free energy that is required for release of ATP come from?

A

proton-motive force (PMF)

250
Q

Structure of F1

(subunit Y

A

a3b3yde

subunit y = **shaft **that passes through Fo & associates with only 1 B subunit

251
Q

each B subunit of F1 has (2)

A

a) 1 catalytic site for ATP synthesis
v) adopts 3 different conformations

252
Q

3 different conformations of each B subunits

A

i) empty (associated with y)
ii) binds ADP
iii) binds ATP

253
Q

Structure of Fo

A

composed of 3 subunits (ab2c10-12)

all transmembrene proteins

mostly alpha-helical

254
Q

In Fo, the membrane embedded cylinder of c subunits is attached to?

A

shaft made up of F1 subunits y & e

255
Q

as protons flow through the membrane from P to N side through Fo, what happens?

A

cylinder & shaft rotate, & B subunits of F1 change conformation as Y subunit associates with each in turn

256
Q

Proposed Rotational Catalysis Mechanism

**(3) **

A

3 active sites of F1 take turns catalyzing ATP synthesis

ADP & Pi are bound -> conformation change

new structure tightly binds ATP

2nd conformational change reduces affinity

257
Q

the second conformational change of the active site is caused by?

A

proton diffusion through Fo, causing c subunites and attached y subunit to rotate.

contact with B subunit forces releae of ATP

258
Q

PMF causes rotation of centrl shaft (y subunit) which comes into contact with each aB subunit pair in succcession.

What happens to:

B-ATP site

B-empty site

B-ADP site

A

cooperative conformational change in which B-ATP site is converted to B-empty conformation and ATP dissociates

B-ADP site is converted to B-ATP conformation, promoting condensation of bound ADP + Pi to form ATP

**B-empty site becomes B-ADP site **which loosely binds **ADP + PI **entering from solvent

259
Q

**malate-aspartate shuttle **used in… for?

A

used in liver, heart & kidney for transporting reducing equivalents from cytosolic NADH into mito matrix

260
Q

**malate-aspartate shuttle **

A

NADH in cytosol enters IMS through outer membrane (porins) passes **2e to OAA forming malate. **

malate crosses IMM and passes 2+ to NAD+ (resulting NADH oxidized by ETC) OA formed cannot pass into cytosol so transaminated to aspartate and leaves via glu-asp transporter

then **OAA is regenerated in cytosol from asp **completing cycle

261
Q

Glyceraldehyde-3-Phosphate Shuttle

A

alternative means of moving reducing equivalents (e-) from cytosol to matrix in skeletal muscle & brain

262
Q

**glycerol-3-phosphate shuttle **

A

in cytosol, DHAP accepts 2 e from NADH (catalyzed by **cytosolic glycerol-3-p DH) **

isozyme of g3p DH bound to outer face of IMM transfer 2e from glycerol-3-p in IMS to FAD then ubiquinone (Q)

263
Q

Regulation of ATP synthesis (ETC & ATP synthase)

A

regulated by availability of ADP

when mass action ratio [ATP]/[ADP][Pi] drops (meaning more ADP available - rate of ATP synthesis increases

264
Q

Regulation of ETC & ATP Synthesis (2)

A

regulation by cellular energy needs

co-ordinate regulation

265
Q

**co-ordinate regulation of oxidative phosphorylation **

A

relative [ATP] & [ADP] controls electron transfer, ox. phos., CAC & glycolysis

266
Q

When ATP consumption increases.. what (3) things happen

A

1) rate of e transfer & ox phos. **increase **
2) rate of pyruvate ox. (via CAC) is enhanced, **increasing flow of e’s **into ETC
3) rate of glycolysis is **enhanced **& supplies more pyruvate

267
Q

what supplements the action of the adenine nucleotide system?

A

interlocking of **glycolysis (cytosol) **and **CAC (mito) **by **citrate **which inhibits **glycolysis **

268
Q

Regulation of **ATP-producing pathways **

when CAC is idling in higher [ATP] .. how does this slow glycolysis?

A

citrate accumulates within mito, then tranported to cytosol

when both [ATP] & [citrate] rise, they produce a **concerted allosteric inhibition of PFK-1 **that is greater than the sum of their individuals effects, slowing glycolysis

269
Q

Regulation of Hexokinase

Glucose –> G6P

A

**product inhibition: **G6P

**activation **by Pi

270
Q

Regulation of Phosphofructokinase-1

F6P –> F-1,6-BP

A

**activated **by: AMP, ADP

**inhibited by: **ATP, citrate

271
Q

regulation of pyruvate kinase

PEP –> pyruvate

A

activated by: ADP

inhibited by: ATP, NADH

272
Q

regulation of PDH complex

A

activated by: ADP, AMP, NAD+

inhibited by: ATP, NADH

273
Q

regulation of citrate synthase

A

product inhibition by citrate

activated by: ADP

inhibited by: ATP, NADH

274
Q

regulation of isocitrate DH

A

activated by: ADP

inhibited by: ATP

275
Q

regulation of a-KG DH

A

product inhibition by **succinyl-CoA **

inhibited by: ATP, NADH

276
Q

The P/O ratio (or the P/2e- ratio)

A

phosphorylation/oxidation ratio

amount of ATP produced from oxygen reduced

amounge of ATP produced from movement of 2e- through ETC

277
Q

explain why 1 NADH produces 2.5 ATP & 1 FADH2 produces 1.5 ATP

A

10 H+ pumped out per NADH

6 H+ pumped out per FADH2

3 H+ flowed in through Fo/F1 per ATP

1H+ used in transporting ATP, ADP, Pi across IMM

10/4 = 2/5 ATP per NADH

6/4 = 1.5 ATP per NADH

278
Q

glycogen

A

major storage of glucose in animals

279
Q

glycogen synthesis takes place in what tissues? predominantly where?

A

takes place in **all tissues **but predominantly in **liver cytosol **

280
Q

Fatty acids cant be converted to glucose in mammals because…

A

cant be catabolized anaerobically

281
Q

Once stored in cytosolic granules, **glycogen can be **(2).

A

1) broken down for distribution to other tissues (liver)
2) broken down for glycolytic fuel to produce **ATP (muscle) **

282
Q

Glycogen structure & characteristics benefits

A

branched to make it more soluble & creating more non-reducing ends that are available for polymerization & breakdown

283
Q

Glycogen synthesis occurs under what type of conditions

A

when [glucose] & [ATP] are high

284
Q

How is glycogen synthesized?

(first step: liver vs muscle)

A

first, glucose is primed by

a) glucokinase (liver)
b) hexokinase (muscle)

285
Q

After glucose is primed by either glucokinase (liver) or hexokinase (muscle) to form G6P…

A

g6p is isomerized by **phosphoglucomutase **

g6p –> g1p

286
Q

After G6P is isomerized by **phosphoglucomutase **to G1P …

A

glucose is charged with UDP by **UDP-glucose pyro-phosphorylase **

sugar-P + NTP => NDP-sugar + 2Pi

287
Q

Strategy of Polymerization

A

sugar nucleotides are suitable for polymerization

Anomeric of sugar activated by attachment to nucleotide through phosphate ester linkage

288
Q

Why attach nucelotide through phosphate ester linkage ? (4)

A

1) net reaction gives large -ve free E change & charges synthetic rxn
2) nucleotide contributes to binding sites for enzymes (glycogenin & glycogen synthase) reducing Ea
3) nucleotidyl group facilitates Nu attack by activating the sugar anomeric C (UDP is good LG, leaving group activation)
4) nucleotides act as tags for different processes (glycogen synthesis vs glycolysis)

289
Q

After glucose is charged with UDP by UDP-glucose pyrophosphate …

A

glucose is transferred to non-reducing end of branched glycogen by **glycogen synthase **

free E change from G1P to glycogen polymer is highly favorable

  • non-reducing end of glycogen acts as Nu and attacks C1 of UDP-glucose, UDP is LG
290
Q

Branching of glycogen

A

block of residues is transferred to make a1->6 linkage from growing a1->4 chain by glycogen branching enzyme

291
Q

Chain & Branchpoints of Glycogen

A

C0hain: a1->4

Branch points: a1-6

once **11 residues **are built up, **6-7 residues **are transferred to a branch

292
Q

Benefits of Branching glycogen

A

increases **solubility & # of non-reducing ends **

293
Q

**glycogenin **catalyzes 2 distinct reactions. (acts as primer)

A

initial attack by OH group of Tyr194on C-1 of glucosyl moiety of UDP-glucose results in glucosylated Tyr residue

C1 of another UDP-glucose molecule is now attacked by C-4 hydroxyl group of terminal glucose and this sequence repeats to form beginning glycogen molecule (primer) of 8 glucose residues attached by **(a1->4) **glycosidic linkages

294
Q

Structure of glycogen particle

A

starting at center of glycogenin molecule, glycogen chains **(12-14 residues) **extend in tiers.

Inner chains have two (a1->6) branches each. Chains in outer tiers are **unbranched. **

12 tiers in mature glycogen particle consisting of **~55,000 glucose residues **

295
Q

**Glycogen Breakdown **by ___ using ___ to form?

A

**glycogen phosphorylase **using **Pi **to form **glucose-1-P **

296
Q

Mechanism of **glycogen phosphorylase **

A

oxygen of Pi attacks anomeric C of non-reducing end forming G1P

process continues until enzyme is 4 glucose units away from branch

297
Q

What happens when glycogen phosphorylase is 4 glucose units away from branch

A

the **glycogen branching enzyme **removes branch

298
Q

Why is Pi not H2O by **glycogen phosphorylase **used to form G1P? (2)

A

1) to keep inside
2) to **save ATP **

299
Q

How is Glycogen broken down?

A

**glycogen phosphorylase **uses Pi to form G1P until 4 glucose units away from branch point

**debranching enzyme **transfers 3 of glucose residues from branch point to chain (transferase activity)

debranching enzyme removes 1 glucose residue of branch point (a1–>6 glucosidase activity)

300
Q

What happens to the G1P formed from glycogen breakdown? (muscle vs liver)

A

**phosphoglucomutase **converts G1P to G6P that

**muscle - **can enter glycolysis

**liver - **converted to glucose by G6Phosphatase for release to blood (GNG)

301
Q

Regulation of **glycogen synthase **and glycogen phosphorylase?

A

reciprocal regulation by **phosphorylation by cAMP dependant pathway **

synthase - less active

phosphorylase - more active

302
Q

Regulation of Glycogen phosphorylase

A

phosphorylase a **phosphatase (PP1) **dephosphorylates - **less active (b) **

phosphorylase b **kinase **phosphorylates - **more active (a) **

303
Q

Phosphorylase:

(a) by b **kinase **(active)
(b) by a **phosphatase **(inactive)
* regulation of each*

A

phosphorylase b kinase activated by **glucagon (liver) **and **epinephrine, **Ca2+, AMP (liver)

304
Q

regulation of glycogen synthase

A

phosphorylated by **GSKinase ** (inactive)

dephos. by **PP1 **(active)

305
Q

regulation of GSK3 (glycogen synthase b)

A

GSK3 phosphorylates to make b conformation

**insulin inhibits GSK3 **

306
Q

regulation of PP1 (glycogen synthase a)

A

PP1 makes glycogen synthase **a conformation **(active)

activated by: **insulin, G6P, glucose (forward activation)

**inhibited by: **glucagon, epinephrine

307
Q

**Epinephrine **targets ___ and **insulin & glucagon **target ___

A

muscle

liver

308
Q

Epinephrine (muscle), insulin & glucaon (liver) regulate…

A

glucose & glycogen synthesis/breakdown

309
Q

epinephrine - release & mechanism

A

adrenaline (mobilizing fuel)

stress leads to fight or flight response

epinephrine released **increases BP, heart beart, **dilation of resp. pathways

increases O2 delivery & uptake in tissues

acts on muscle, adipose, liver in cAMP dependant pathway that **inactivates **glycogen synthase and **activates **glycogen phosphorylase (increasing glucose release)

310
Q

Signal Transduction Pathway of Epinephrine (6)

A

1) binds

2) conformational change -> GTP binds Gsa subunit (replaces GDP)
3) Gsa associates with **adenyl cyclase **
4) cAMP formation (catalyzed by adenyl cyclase)
5) activation of PKA
6) phosphorylation of other proteins (kinases) by PKA

311
Q

Glucagon & Insulin are triggered by?

A

blood glucose levels

312
Q

Glucagon acts like ___ (via ___ ) to promote? inhibit? while stimulating?

A

epinephrine regulation (via **cAMP) **to promote **glycogen breakdown **& inhibit **glycolysis **while stimulating **GNG & glucose release (liver) **

313
Q

Insulin promotes?

A

glucose uptake & storage/consumption

314
Q

why do you think regulation of fatty acid and carbohydrate metabolism involves **hormones **but amino acid & nucleic acid metabolism doesnt?

A

FAs & carbs make up **main storage **fuels in adipocytes & muscle, respectively

aa’s & nucleic acids contribute to functional & informational macromolecules

315
Q

Hormone

A

a chemical released by a cell or gland in one part of the body that sends out messages that affects cells in other parts of the organism

316
Q

The only source of energy that the body can use…

A

ATP

stored E (fat, glycogen, creatine phosphate) must first be converted to ATP before body can actually use it

Bioc 221 (4 decks)

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