MCB 102- Midterm 2

Thomas

1st Law of Thermodynamics

energy can not be created or destroyed

2nd Law of Thermodynamics

entropy of the universe increases with every spontaneous reaction

___ pathways converge and anabolic pathways _____

catabolic, diverge

ΔG° > 0

endergonic, Keq < 1, prefers reactants

ΔG° < 0

exergonic, Keq > 1, prefers products

what reaction is this?

aldol condensation

what reaction is this?

Claisen Condensation

what reaction is this?

decarboxylation

ATP hydrolysis

Dependent on Mg2+

free energy change is large and negative

products of ATP are more stable: they repel each other and Pi is stabilized by resonance delocalization

How does ATP provide energy?

via group transfers

what group in ATP is the leaving group

gamma-phosphate

oxidation is ____ of electrons

reduction is ____ of electrons

loss; gain

if something is a reducing agent it is being ____

oxidized

first source of electrons in glycolysis

reduced carbon

final acceptor in glycolysis

O2

how are all half-reactions listed

as reductions

what E value is reduced v. oxidized

higher E is reduced, lower E is oxidized

E (more positive) - E (more negative)

what does high ΔE mean

redox reaction is favorable

how are e- transferred between biomolecules

1. directly as electrons

2. as hydrogen atoms

3. as hydride ion

   1. through combination with oxygen

HEEP

high energy electron pairs

NAD+, NADP, FMN, FAD

what cofactor is this?

NAD+/NADH, NADP+/NADH

what cofactor is this?

FAD → FADH (vitamin B2)

what cofactor is this?

Niacin (vitamin B3)

steps of catabolism

1. glycolysis

2. citric acid cycle

3. oxidative phosphorylation

where does glycolysis occur?

cytoplasm/cytosol

net glycolysis equation

glycolysis + 2 NAD+ + 2 ADP + 2 Pi → 2 pyruvate + 2 NADH + 2 ATP + 2 H+ + 2 H20

glycolysis: step 1 (first priming reaction)

glucose → glucose 6-phosphate

hexokinase

ATP required

adding phosphate to oxygen of C6

irreversible step

glycolysis: step 2

glucose 6-phosphate → fructose 6-phosphate

phosphohexose isomerase

5 carbon ring to 4 carbon ring (carbon 1 becomes substituent of carbon 2)

glycolysis: step 3 (second priming reaction)

fructose 6-phosphate → fructose 1,6-biphosphate

ATP required

phosphofructokinase-1 (PFK-1)

add Pi to oxygen of C1

irreversible step

allosterically inhibited by ATP; activated by AMP/ADP

glycolysis: step 4

fructose 1,6-biphosphate → glyceraldehyde 3-phosphate (G3P)

adolase

cleavage of 6-carbon sugar to 2 3-carbon sugar phosphates (reverse aldol reaction)

glycolysis: step 5

dihydroxyacetone phosphate → glyceraldehyde 3-phosphate

triose phosphate isomerase

right sugar phosphate with ketone needs to be moved around to become aldehyde (2 of the products now)

glycolysis: step 6 (start of payoff phase)

2 glyceraldehyde 3-phosphate + 2 Pi + 2 NAD+→ 1,3-bisphosphoglycerate + 2 NADH + 2 H+

glyceraldehyde 3-phosphatase dehydrogenase: active site of cysteine forms high-energy thioester intermediate

oxidation and phosphorylation

replaces hydrogen (goes to NADH) with oxygen + phosphate at carbonyl carbon

glycolysis: step 7

2 1,3-bisphosphoglycerate + 2 ADP → 3-phosphoglycerate + 2 ATP

phosphoglycerate kinase

substrate-level phosphorylation (removing phosphate from carbonyl oxygen)

glycolysis: step 8

2 3-phosphoglycerate → 2 2-phosphoglycerate

phosphoglycerate mutase

moves phosphate from carbon 3 to carbon 2

glycolysis: step 9

2 2-phosphoglycerate → 2 phosphoenolpyruvate + 2 H2O

enolase

removes OH group from end carbon (dehydration)

glycolysis: step 10

2 phosphoenolpyruvate + 2 ADP → 2 pyruvate + 2 ATP

pyruvate kinase

substrate-level phosphorylation (remove phosphate from oxygen and turn it into double-bonded oxygen)

irreversible step

how is glycolysis favorable?

some of the intermediate steps of glycolysis are unfavorable, but some are very favorable making the net reaction favorable

two fates of pyruvate

fermentation: ethanol or lactic acid

oxidation: acetyl-CoA

what happens to all the HEEPs from glycolysis in fermentation

Le Chat’s Principle: the concentration of NADH is too high which results in it being used to create NAD+

lactic acid fermentation

2 pyruvate + 2 NADH → 2 lactate + 2 NAD+ + 2 H+

lactate dehydrogenase

turns carbonyl into alcohol

seen in active muscle and lactic acid bacteria

why is pyruvate being reduced and not oxidized?

purpose of reaction and no final O2 acceptor in fermentation

ethanol fermentation: step 1

2 pyruvate → 2 acetaldehyde + 2 CO2

pyruvate decarboxylase

removes CO2 and creates aldehyde

TPP as a cofactor

ethanol fermentation: step 2

2 acetaldehyde + 2 NADH + 2 H+→ 2 ethanol + 2 NAD+

alcohol dehydrogenase

reduction reaction

get rid of double bond and give hydrogen to O and carbonyl C

cofactor: Zn2+ (polarized aldehyde oyxgen)

what coenzyme is this

TPP (vitamin B1)

TPP mechanism

stabilizes carbanion intermediates during decarboxylation of pyruvate

3 stages of cellular respiration and where do they occur

1. Acetyl-CoA production (pyruvate oxidation), matrix

2. Citric Acid Cycle (acetyl-coA oxidation), matrix

3. Oxidative Phosphorylation (HEEP carrier oxidation), inner membrane

what coenzyme is this?

coenzyme A (vitamin B5)

what coenzyme is this?

lipoyllysine (lipoate)

net reaction of acetyl-CoA production

pyruvate + CoA-SH + NAD+ + TPP + lipoate + FAD → acetyl-CoA + NADH + CO2

pyruvate dehydrogenase complex (E1 + E2 + E3)

inhibited by products and ATP, activated by reactants and AMP

pyruvate dehydrogenase complex enzymes

pyruvate dehydrogenase, dihydrolipoyl transacetylase, dihydrolipoyl dehydrogenase

acetyl-CoA production: step 1

pyruvate + TPP → CO2 + hydroxyethyl-TPP (pyruvate decarboxylation)

pyruvate dehydrogenase

TPP attacks carbonyl group and kicks off CO2

acetyl-CoA production: step 2

transfering acetyl group to enzyme 2

TPP is regenerated

hydroxyethyl-TPP + lipoic acid → acetyl lipoyllysine + TPP

acetyl-CoA production: step 3

actually producing Acetyl-CoA

dihydrolipoyl transacetylase (E2)

acetyl lipoyllysine + CoA-SH → acetyl-CoA + reduced lipoyllysine

removing lipoic acid and adding CoA-SH to acetyl group

acetyl-CoA production: steps 4-5

dihydrolipoyl dehydrogenase (E3)

transfer 2 hydrogens from reduced lypoyl groups to FAD

FADH2 gets oxidized by NAD+ turning it into NADH + H+

net reaction of citric acid cycle

acetyl-CoA + 3 NAD+ + FAD + GDP + Pi + 2 H2 O → CoA-SH + 3 NADH + FADH2 + 3 H+ + GTP + 2 CO2

what is the goal and logic of CAC?

oxidize acety-coA as much as possible

combine acetyl-coA with 4 carbon molecule making it easier to oxidize

CAC: step 1

acetyl-CoA + H2O + oxaloacetate→ citrate + CoA-SH

citrate synthase

aldol condensation (methyl group attacks carbonyl on oxaloacetate)

inhibited by NADH, succinyl-CoA, citrate, and ATP; activated by ADP

what product of citric acid cycle inhibits glycolysis and why

citrate inhibits PFK-1 to keep glycolysis and CAC in sync

what is weird about citrate synthase

it needs precise positioning of the active site

CAC: step 2

citrate → H2O + cis-Aconitate → isocitrate

aconitase

dehydration and then rehydration (OH group is respositioned because secondary alcohol is good substrate for oxidation)

CAC: step 3

isocitrate + NAD+ → a-ketoglutarate + NADH + H+

isocitrate dehydrogenase

oxidative decarboxylation (COO- leaves and double on adjacent carbon is formed by oxygen)

NAD (1/3)

irreversible step

inhibited by ATP; activated by Ca2+ and ADP

CAC: step 4

a-ketoglutarate + CoA-SH → succinyl-CoA + CO2

a-ketoglutarate dehydrogenase complex

oxidative decarboxylation (TPP leads to decarboxylation and CoA-SH performs acyl sub)

similar to pyruvate-dehydrogenase

NAD (2/3)

inhibited by succinyl-CoA and NADH; activated by Ca2+

CAC: step 5

succinyl-CoA + GDP + Pi → succinate + GTP + CoA-SH

succinyl-CoA synthetase

substrate-level phosphorylation (thioester is kicked off and that energy goes to GTP)

CAC: step 6

succinate + FAD+ → fumarate + FADH2

succinate dehydrogenase

dehydrogenation (double bound between two CH2s is created kicking off 2 Hs)

CAC: step 7

fumarate + H2O → malate

fumarase

hydration (add water and removed CH-CH double bound)

CAC: step 8

malate → oxaloacetate

malate dehydrogenase

dehydrogenation (remove hydrogen from oxygen and make double bound)

NAD (3/3)

low [oxaloacetate] can help make this reaction more favorable

which CAC intermediates can be used in anabolism?

citrate → fatty acids

a-ketoglutarate, oxaloacetate → amino acids

oxaloacetate → gluconeogenesis

net equation for gluconeogenesis

2 pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 6 H2O → glucose + 4 ADP + 2 GDP + 6 Pi + 2 NAD+

anabolic pathway (build molecules, requires ATP)

kidneys and livers perform gluconeogenesis

what enzymes are used in glucogenesis to bypass irreversible steps

glucose 6-phosphatase instead of hexokinase

fructose 1,6-bisphosphatase-1 instead of PFK1

pyruvate carboxylase and PEP carboxylase instead of pyruvate kinase

gluconeogenesis: 1st bypass step 1

bicarbonate + pyruvate + ATP → oxaloacetate + ADP + Pi

cofactor: biotin (activates CO2 to leave and add to pyruvate)

pyruvate carboxylase (attach CO2 to methyl end of pyruvate)

malate-aspartate shuttle (used in step 1 of gluconeogenesis)

oxaloacetate cannot cross membrane to get to cytoplasm

1. oxaloacetate + NADH → malate + NAD+

2. malate + NAD+ → oxaloacetate + NADH

3. oxaloacetate + glutamate → aspartate + a-ketoglutarate

4. aspartate + a-ketoglutarate → oxaloacetate + glutamate

gluconeogenesis: 1st bypass step 2

oxaloacetate + GTP → phosphoenolpyruvate + GDP + CO2

PEP carboxykinase (remove CO2, make enolate, add phosphate to carbonyl)

gluconeogenesis: bypass 2

fructose 1,6-bisphosphate + H2O → fructose 6-phosphate + Pi

fructose 1,6-bisphosphatase-1 (adds water, removes phosphate)

inhibited by AMP

gluconeogenesis: bypass 3

glucose 6-phosphate + H2O → glucose + Pi

glucose 6-phosphatase (adds water, removes phosphate)

how much more ATP and GTP is used in gluconeogenesis and why

2 of each; because we are reversing glycolysis and trying to overcome lots of irreversible steps

what hormone starts glycolysis

insulinwhat h

what hormone starts gluconeogenesis

glucagon

isoenzymatic regulation: difference between hexokinase 1 and hexokinase 4

1 is expressed in most tissues (low Km, inhibited by GCP product)

4 is expressed in liver (high Km, ensures that it only stores glucose when its plentiful)

CAC is an _____ pathway

amphibolic (can be catabolic or anabolic)

how are CAC intermediates constantly replenished

via anaplerotic reactions: pyruvate carboxylase replenishes oxaloacetate

what are the regulated steps of CAC

step 0, 1, 3, 4

what are the two parts of oxidative phosphorylation?

ETC and ATP Synthase

3 important coenzymes in ETC

NADH

FADH2

Ubiquinone (can take 1 or 2 e-)

2 metalloproteins in ETC

cytochromes: carry heme prosthetic groups

Fe-S: coordinated by cysteine residues

both are 1e- acceptor/donorsn

net ETC equation

2NADH + 22H+ + O2 → 2NAD+ + 20H+ + 2H20ETC:

ETC: step 1

NADH donates 2 electrons to complex 1 (NADH dehydrogenase)

electrons go to Q → QH2 (reduced) (go to Fe-S cluster before)

4 H+ pumped out

ETC: step 2

complex 2 (succinate dehydrogenase), also part of CAC

FADH2 donates 2 electrons to Q → QH2 (electrons go to Fe-S cluster before)

FAD gets its electrons from succinate → fumarate in CAC

no protons pumped

ETC: step 3

complex 3 (medley of iron centers)

QH2 transfers its electrons to complex 3

e- move to cyt-c (1 at a time)

4 H+ are pumped out

ETC: step 4

e- are transferred to complex 4

O2 is the final electron acceptor

2 H+ are pumped out

what is the chemiosmotic theory

1. ETC creates proton gradient (more H+ outside)

2. accumulation of H+ outside creates an electrochemical gradient which stores energy as proton motive force

3. protons flow back into matrix via ATP synthase

   1. ATP synthase rotates leading to creation of ATP

ATP synthase structure

F1: in the matrix, catalyzes ATP synthesis in beta subunit

F0: transports protons from intermembrane space to matrix, c ring spins, energy transferred to F1 to catalyze ADP phosphorylation

c ring spins (bc of protons) and wacks beta unit up → changes shape of beta → this force stimulates ATP synthesis

3 conformations of beta subunit of F1

B-ADP = binds ADP + Pi loosely

B-ATP = binds ATP tightly

B-empty = loose-binding

how many ATP from 1 spin of ATP synthase

3 ATP

how much ATP per NADH and FADH2

2.5 ATP

1.5 ATP

total ATP yield

7 ATP (glycolysis) + 5 ATP (pyruvate oxidation) + 20 ATP (citric acid) = 32 ATP

why are fats great for long-term energy storage

1. lots of reduced carbons in CH2 (oxidative potential)

2. pack molecules tightly

3. very stable

4. long term storage

5. slow delivery

fatty acid breakdown steps

1. make fatty acid-CoA

2. transport from cytosol to mitochondria

3. beta oxidation (break down 2C’s at a time)

fatty acid breakdown: step 1

activate FA by attaching CoA

consumes 2 ATP for activation

dehydrogenation → hydration → dehydrogenation → thiolysis

fatty acid breakdown: step 3

beta-oxidation

similar to citric acid cycle step

1. dehydrogenation with FAD (lose 2 H+)

2. hydration

3. dehydrogenation with NAD+

1 round of b-oxidation makes how many acetyl-CoA, NADH, and FADH2

1 acetyl-CoA, 1 NADH, 1 FADH2

for N carbon fatty acid → N/2 acetyl CoA, (N/2) - 1 NADH, FADH2