Lecture 2 Topics

Finish discussion of thermodynamics (G, E)

ATP as an universal carrier of chemical energy
Role of enzymes and co-factors

Summary of Lecture 1
Need for metabolism
Provides building blocks for regeneration
Energy conversion compatible with C-based life

Chemical bonds as stores of energy (H)

Rearrangement of bonds release or require H
(- H, exothermic; + H, endothermic)

Free Gibbs Energy (G)

G = H TS
G = Go + RTlnQ
G = nFE

Review

 Reduction Potential (E)

yet another way to express G

Metabolic reactions are often redox reactions, involving

transfer of electrons from a donor to an acceptor

1. Direct combination with oxygen (X X + O=O 2O

X)

2. Transfer of the hydride anion (HH, or H- + H+)

3. Transfer of hydrogen (H, or e- + H+)
4. Direct transfer (e-)

Lecture 1

 Redox reactions can be written as two half-reactions

(by convention:
convention each is written in the direction of the reduction!)
reduction

Aoxidized + Breduced Areduced + Boxidized

1. Aoxidized + e- Areduced
2. Boxidized + e- Breduced

Electronegativities (Tab. 1) can predict direction of e- transfer

See p. 6

Lecture 1

Question: Which half-reaction has the higher

affinity for electrons at standard conditions ?
Reference
Electrode:

H+ + eH2 half-reaction (1M each)

(Eo= 0.00 V)

Test
Electrode:

A+ + eA
(Eo= ??? V)

half-reaction (1M each)

By convention:
convention
If e- flow from reference to test (test is stronger e- acceptor): Eo > 0 (+V)
If e- flow from test to reference (test is weaker e- acceptor):

See p. 6

Eo < 0 (- V)

Lecture 1

Table 3: Standard Reduction Potentials for Biological Reduction HalfHalf-Reactions

Reactions
Half-Reaction
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
36.
37.
38.

Eo(V)

(written as reduction)

Excited(Chlorophyll)2*
Succinate + CO2 + 2H+ + 2e- -Ketoglutarate + H2O
Acetate + 2H+ + 2e- Acetaldehyde + H2O
SO42- + 2H+ + 2e- SO32- + H2O
Ferredoxin (Fe3+) + e- Ferredoxin (Fe2+)
2H+ + 2e- H2 (at pH 7.0)
CO2 + H+ + 2e- Formate
-Ketoglutarate + CO2 + 2H+ + 2e- Isocitrate
Acetoacetate + 2H+ + 2e- -Hydroxybutyrate
Cystine + 2H+ + 2e- 2 Cysteine
NADP+ + 2H+ + 2e- NADPH + H+
NAD+ + 2H+ + 2e- NADH + H+
Lipoic acid + 2H+ + 2e- Dihydrolipoic acid
1,3-bisPGA + 2H+ + 2e- 3-PGA + Pi
S + 2H+ + 2e- H2S
Glutathione + 2H+ + 2e- 2 Glutathione (reduced)
FAD + 2H+ + 2e- FADH2 (Flavoprotein 0.003-0.091)
FMN + 2H+ + 2e- FMNH2
Acetaldehyde + 2H+ + 2e- Ethanol
Pyruvate + 2H+ + 2e- Lactate
Oxaloacetate + 2H+ + 2e- Malate
Crotonyl-CoA + 2H+ + 2e- Butyryl-CoA
2H+ + 2e- H2 (at standard conditions, 1 M = pH 0)
Fumarate2- + 2H+ + 2e- Succinate2UQ + 2H+ + 2e- UQH2
Cytochrome b (Fe3+)+ e- Cytochrome b (Fe2+)
Cytochrome c1 (Fe3+)+ e- Cytochrome c1 (Fe2+)
Cytochrome c (Fe3+)+ e- Cytochrome c (Fe2+)
Rieske Fe-S (Fe3+)+ e- Rieske Fe-S (Fe2+)
Cytochrome a (Fe3+)+ e- Cytochrome a (Fe2+)
O2 + 2H+ + 2e- H2O2
Cytochrome a3 (Fe3+)+ e- Cytochrome a3 (Fe2+)
Cytochrome f (Fe3+)+ e- Cytochrome f (Fe2+)
NO3- + 2H+ + 2e- NO2- + H20
Photosystem P700
Fe3+ + e- Fe2+
O2 + 4H+ + 4e- 2H20
(Chlorophyll a)2+ + e- (Chlorophyll a)2

+
+
+
+
+
+
+
+
+
+
+
+
+
+
+

1.0
0.67
0.58
0.57
0.43
0.42
0.42
0.38
0.35
0.34
0.32
0.32
0.29
0.29
0.24
0.23
0.22
0.22
0.20
0.18
0.17
0.02
0.00
0.03
0.06
0.08
0.22
0.25
0.28
0.29
0.30
0.35
0.37
0.42
0.43
0.77
0.82
1.10

Direction of
spontaneous
electron flow.

See p. 3

Reduction Potential E for each Half-Reaction (Nernst Equation)

1. Aoxidized +

e-

Areduced

2. Boxidized +

e-

Breduced

E=

Eo

RT [e- Acceptor, Aox]

+ nF ln [e- Donor, Ared]

E=

Eo

RT [e- Acceptor, Box]

+ nF ln [e- Donor, Bred]

E of Redox Reaction (Aox + Bred Ared + Box)

E = EOxidant (A) EReductant (B)
Relationship between E and G
G = nFE

Go = nFEo

E = G/nF

Eo = Go/nF

See p. 6

Table 3: Standard Reduction Potentials

Half-reaction (written as reduction by convention)
Excited (Chlorophyll a)2*

Eo (V)
~ - 1.00

Acetate + 2H+ + 2e- Acetaldehyde + H2O

- 0.58

NAD+ + 2H+ + 2e- NADH + H+

- 0.32

Pyruvate + 2H+ + 2e- Lactate

- 0.18

2H+ + 2e- H2 (at standard conditions, 1M each, pH 0)

0.00

NO3- + 2H+ +2e- NO2- + H2O

+ 0.42

O2 + 4H+ + 4e- 2H2O

+ 0.82

(Chlorophyll a)2.+ + e- (Chlorophyll a)2

+ 1.10
e- flow

See p. 3

Metabolic Life Styles

Chemolithotrophs

Chemoorganotrophs

Fully or partially reduced

inorganic compounds
(e.g., H2, NH3, NO2-,
H2S, S2O32-, S, Fe2+)

Organic compounds
(e.g., sugars, amino or
fatty acids, organic
acids, etc.)

e-

ERed

Initial Electron Donor

E = EOx ERed

Energy Metabolism

E = G/nF

Terminal Electron Acceptor

(e.g., NO3-, NO2-, SO42-, Fe3+,
CO2, partially oxidized
organic compounds)
Anaerobes

O2

e-

EOx

Aerobes

p. 7

Organic
Compound

Eo

Oxidized
Compound

( - 0.60 V)

(+ 0.42 V)

Nitrate (NO3-)

Nitrite (NO2-)
(+ 0.42 V)

(+ 0.82 V)

H 2O

1/2 O2

p. 7

Food

CO2

H 2O

O2

Reduced
Carbon

Respiration

ENERGY

Humans and Animals

Heterotrophic Metabolism

p. 8

Plants and
Photosynthetic Bacteria

Autotrophic Metabolism

LIGHT

Photosynthesis

Fossil Fuels

Day

CO2

H2 O

O2

Reduced
Carbon

Night

Respiration

ENERGY

Food

p. 8

Plants and
Photosynthetic Bacteria

Autotrophic Metabolism

LIGHT

Fossil Fuels

Photosynthesis
Day

CO2

H2O

O2

Reduced
Carbon

Night

Respiration

Food

ENERGY

CO2

H2O

O2

Reduced
Carbon

Respiration

ENERGY

Humans and Animals

Heterotrophic Metabolism

p. 8

JACOB VAN RUYSDAEL (1652)

2H2O

2{H2} + O2
2e- + 2H+
(0.7V)

ATP

O2 (+0.8V)

H2O

Hydrolysis Reactions of Phosphate Esters and Anhydrides

Phosphate esters
O

O
R O

P
O-

O-

H2O

ROH + HO

O-

O-

p. 9

Phosphate anyhydride
O
R O

P
O-

O
O

P
O-

O
O- + H2O

R O

P
O-

OH + HO

O-

O-

p. 9

Acyl phosphate

C O

P
O-

O
O- + H2O

OH + HO

O-

O-

p. 9

ATP (Adenosinetriphosphate), ADP, and Their Mg2+ Complexes

Phosphoester bond
NH2

Phosphoanhydride
bonds

O
-O

P
O-

O
O

P
-

P
O

(Adenine)

(Ribose)
H

OH

H
OH

Mg2+

MgATP
p. 10

NH2
Phosphoanhydride
bond

O
-O

O
O

P
-

OH

OH

H
OH

Mg2+

MgADP

p. 10

The ATP Gun

Spring-loaded phosphate bullets

The gun is safe (kinetically stable),

until you pull the trigger (to overcome activation energy)

Cause for G
of Hydrolysis
o

G
(kJ/mol)

Transfer
Potential

Phosphoenolpyruvate
(Pyruvate + Pi)

- 62.2

62.2

Enolic
phosphate

Tautomerization of product (Pyr);

Resonance stability of Pi

1,3-Bisphosphoglycerate
(3-PGA + Pi)

- 49.6

49.6

Acyl
phosphate

Ionization of product (3-PGA);

Resonance stability (Pi, 3-PGA)

Phosphocreatine
(Creatine + Pi)

- 43.3

43.3

Guanidine
phosphate

Resonance stability of product

(creatine)

Pyrophosphate (PPi)
(Pi + Pi)

- 33.6

33.6

Phosphoric acid
anhydride

Electrostatic bond strain in PPi

substrate; Ionization and
resonance stability of Pi group

ATP (ADP + Pi)

- 30.5

30.5

Same as PPi

Same as PPi

ADP (AMP + Pi)

- 30.5

30.5

Same as PPi

Same as PPi

Acetyl-CoA
(and other thioesters)

- 31.5

31.5

Thioester

No resonance stabilization of
Acetyl-CoA;
Ionization and resonance
stabilization of acetate

Glucose-1-P (Glucose + Pi)

- 20.7

20.7

Phosphate
semiacetal

Bonds in glucose-1-P not

that strained

Glucose-6-P (Glucose + Pi)

- 13.9

13.9

Phosphate
ester

Bonds in glucose-6-P not

strained

AMP (Adenosine + Pi)

- 9.2

9.2

Phosphate
ester

Bonds in AMP not strained;

Adenosine does not ionize

0.0

0.0

Phosphate

Compound
(hydrolyzed to)

Type of
Compound

(Acetate + CoA-SH)

Phosphate (Pi)

Table 4: Standard Free

Energies of Hydrolysis

Compounds with
decreasing Go of hydrolysis

p. 4

Cause for G
of Hydrolysis
o

G
(kJ/mol)

Transfer
Potential

Phosphoenolpyruvate
(Pyruvate + Pi)

- 62.2

62.2

Enolic
phosphate

Tautomerization of product (Pyr);

Resonance stability of Pi

1,3-Bisphosphoglycerate
(3-PGA + Pi)

- 49.6

49.6

Acyl
phosphate

Ionization of product (3-PGA);

Resonance stability (Pi, 3-PGA)

Phosphocreatine
(Creatine + Pi)

- 43.3

43.3

Guanidine
phosphate

Resonance stability of product

(creatine)

Pyrophosphate (PPi)
(Pi + Pi)

- 33.6

33.6

Phosphoric acid
anhydride

Electrostatic bond strain in PPi

substrate; Ionization and
resonance stability of Pi group

ATP (ADP + Pi)

- 30.5

30.5

Same as PPi

Same as PPi

ADP (AMP + Pi)

- 30.5

30.5

Same as PPi

Same as PPi

Acetyl-CoA
(and other thioesters)

- 31.5

31.5

Thioester

No resonance stabilization of
Acetyl-CoA;
Ionization and resonance
stabilization of acetate

Glucose-1-P (Glucose + Pi)

- 20.7

20.7

Phosphate
semiacetal

Bonds in glucose-1-P not

that strained

Glucose-6-P (Glucose + Pi)

- 13.9

13.9

Phosphate
ester

Bonds in glucose-6-P not

strained

AMP (Adenosine + Pi)

- 9.2

9.2

Phosphate
ester

Bonds in AMP not strained;

Adenosine does not ionize

0.0

0.0

Phosphate

Compound
(hydrolyzed to)

Type of
Compound

(Acetate + CoA-SH)

Phosphate (Pi)

Phosphoryl-group
transfer potentials

Receive P~group

Donate P~group

p. 4

High Energy
Substrates

Cellular
Macromolecules

ATP + H2O
G

+G

+G

ADP + Pi
Intermediates
of Metabolism
Low Energy
Products

Enzymes (biological catalysts)

do NOT change G of chemical reactions !!!
but decrease their activation energies
inrease the rate (107 to 1019-fold) of attaining
equilibrium (G is not a kinetic constant!)
reaction rates (flux) can be regulated
provide specificity and couple reactions
coordinate many reactions into metabolic
networks (pathways) via shared intermediates

A
Large -G

F
Small -G

OOC
NH3

The 20 Protein Amino Acids

H

L-Amino Acid

(constituents of enzymes)

A. Nonpolar, Aliphatic R-Groups

O
H2N

CH C

O
OH

H2N

CH C

OH

H2N

CH CH3

CH3

H2N

CH C

CH3

Valine
Val, V

O
OH

Leucine
Leu, L
O

OH

CH2
CH2

OH

CH2

CH CH3

Glycine
Gly, G

CH C

HN

H2N

CH

OH

CH

CH3

CH2

S
CH3

Methionine
Met, M

Proline
Pro, P

CH3

Isoleucine
Iso, I

p. 12

B. Aromatic R-Groups

O
H2N

CH C

O
OH

H2N

CH C

O
OH

CH2

CH2

H2N

CH C

OH

CH2

HN

Tryptophan
Trp, W

OH

Tyrosine
Tyr, Y

Phenylalanine
Phe, F

p. 12

C. Polar, Uncharged R-Groups

O
H 2N

CH C

OH

H2 N

CH C

CH 2

CH OH

SH

CH 3

OH

CH C

OH

CH2
O

NH2

Threonine
Thr, T

H 2N

Cysteine
Cys,C

H 2N

Asparagine
Asn, N

CH C OH

H 2N

CH C

OH

CH2

CH 2

CH2

OH

NH2

Serine
Ser, S

Glutamine
Gln, Q

p. 13

D. Positively Charged R-Groups

O
H2N

CH

O
H2N

OH

CH C

OH

H2N

CH2

CH2

CH2

CH2

CH2

NH
NH

Histidine
His, H

OH

CH2

CH2

CH C

NH

NH2

CH2

Arginine
Arg R

NH2

Lysine
Lys, K

E. Negatively Charged R-Groups

O
H2N

CH C

O
OH

CH2

CH2
C

H2N CH C OH
CH2

C O

OH

Aspartate
Asp, D

OH

Glutamate
Glu, E

p. 13

F. Peptide Bond

R1
H 3N

R2

G. Isopeptide Bond

R1
COO-

H3N

NH 3

(CH 2)4

COO-

Lysine
H
H3N

O
CH 2

CH 2

COO-

R2
N

COO-

Glutamate

p. 14

Polar and charged side-chain terminal groups and

carboxylate-arginine and carboxylate-carboxylate dyads

Gutteridge and Thornton (2005) Understanding natures catalytic toolkit.

Trends Biochem Sci 30:622-629

pKa values of polar and charged amino acids

Gutteridge and Thornton (2005) Understanding natures catalytic toolkit.

Trends Biochem Sci 30:622-629

Gutteridge and Thornton (2005) Understanding natures catalytic toolkit.

Trends Biochem Sci 30:622-629

CO-FACTORS
(Non-protein moieties required for catalytic activity)

1. Metals

Structural role (e.g., Mg2+)

Catalytic role (e.g., Fe2+)

2. Co-enzymes

Organic molecules (catalytic)

Co-substrates

- If transiently bound (e.g., ATP)

Prosthetic group - If covalently bound (e.g., FAD)

Must be regenerated if altered in reaction (by same or different enzyme)
Most water-soluble vitamins are precursors of co-enzymes

see p. 14/15