Introduction to metabolismBiochemistry, 4th edition, RH Garrett & CM Grisham, Brooks/Cole (Cengage); Boston, MA: 2010

pp 511-534

Instructor: Kirill Popov

1. Metabolism and metabolic pathways

2. Biochemistry of phosphate compounds

3. Oxidation-reduction reactions

4. Vitamins and co-enzymes

Photosynthesis in plants,algae, bacteria

CO2

Cellular respiration inanimals, plants, algae, bacteria

Reduced fuelsand O2

Cycling of carbon dioxide and oxygen

Amino acidsand otherreduced

nitrogen-carboncompounds

N2NitrateNO3

−Ammonia

NH4+

NitriteNO2

−

nitrogen fixationby some bacteria

(e.g., Azotobacter, Rhizobium,Klebsiella)

degradationby animals andmicroorganisms

synthesis inplants and

microorganisms

nitrificationby soil bacteria

(e.g., Nitrobacter)

denitrification

reductionby some

anaerobicbacteria,

mostplants

Cycling of nitrogen in the biosphere

Cellmacromolecules

ProteinsPolysaccharidesLipidsNucleic acids

Energydepleted

end productsCO2H2ONH3

Catabolism

Anabolism

Chemicalenergy

Energy-containing nutrients

CarbohydratesFatsProteins

Precursormolecules

Amino acidsSugarsFatty acidsNitrogen bases

ADP+PiNAD+

NADP+

FAD

ATPNADH

NADPHFADH2

Relationship between catabolic and anabolic pathways

Chemistry of ATP

Adenine

Ester

AnhydrideAnhydride

Acetic anhydride,a caboxylic acid

anhydride

ATP

Methyl acetate,a caboxylic acid

ester

P O P O P O CH2-O

O

OHOH

O- O- O-

O O O

CH3 C O C CH3

OOCH3 C O

OCH3

ATP4- + H2O → ADP3- + P2-i + H+

ΔG' � = −30.5 kJ/mol

O:H

H

resonance stabilization

O P O P O P OO O O

O O ORib Adenine

−−

−

−

Rib AdenineP O P OO O

O OOH

−−O P

O

OOH

−

−

Rib AdenineP O P OO O

O OO

−

−

−

3−δ−

δ−

δ−

δ−O P OO

OH+ +H+

hydrolysis, withrelief of charge repulsion

ionization

ATP4-

ADP2-Pi

ADP3-

Chemical basis for the large free-energy change associated with ATP hydrolysis

Reaction coordinate

Free

ene

rgy,

G

Reaction 1:glucose + Pi →

glucose 6-phosphate

Reaction 3:glucose + ATP →

glucose 6-phosphate + ADP

Reaction 2:ATP → ADP + Pi

ΔG1

ΔG2 ΔG3

ΔG3 = ΔG1 + ΔG2

Energy coupling in chemical processes

COOHCHCH2

H2N

CH2COHO

+ NH3

NH3

Pi

Glutamine

ATP

ATP

ADP + Pi

ADP

Glutamate

Enzyme-boundglutamyl phosphate

Written as a one-step reaction

Actual two-step reaction

COOHCHCH2

H2N

CH2CNH2O

COOHCHCH2

H2N

CH2COO P

OOHHO

ATP hydrolysis in two steps

-70

1,3-BisphosphoglyceratePhosphocreatine

ATPHigh-energycompounds

Low-energycompounds

ΔG’°

of h

ydro

lysis

(kJ/

mol

)

-60

-50

-40

-30

-20

-10

0

Phosphoenolpyruvate

Pi

Glycerol- PGlucose 6- P

CCHOHOO

CH2 O

P

P P Creatine

COOHCCH2

O P

Adenine Rib PP P

Ranking of biological phosphate compounds by standard free energies of hydrolysis

Alkane

CH2 CH2OH

CH2 CH3

O C O

CCH2HO

O

CH2

O

HC

Alcohol

Aldehyde

Carboxylic acid

Carbon dioxide

The oxidation states of carbon in biomolecules

CH CHO

OCH3

OH O

C CHO

OCH3

Lactate Pyruvatelactate

dehydrogenase

2H+ + 2e−

2H+ + 2e−

An oxidation-reduction reaction

N

HC

O

NH2

CH2O

OH OH

O

PO O-

O

O

OHOH

CH2

N

N

N

N

NH2PO O-

O

+N

C

R

H H O

NH2..

2e-

2H+

NADH(reduced)

NAD+

(oxidized)

adenine

+ H+

In NADP+ this hydroxyl groupis esterified with phosphate

NAD and NADP

N

N N

NH

O

OCH2

COHCOHCOHCH2

OPOPO

O

OHOH

CH2

N

N

N

N

NH2

O

O

-O

-O

CH3

CH3

HHH

N

N N

NH

O-

ROCH3

CH3

HN

N N

NH

R

O

OCH3

CH3

H

H

Isoalloxazine ring

FAD

FMN

H+ + e- H+ + e-

FADH• (FMNH•)(semiquinnone)

FADH2 (FMNH2)(fully reduced)

Flavin adenine dinucleotide (FAD) andFlavin mononucleotide (FMH)

adenine

Structures of oxidized and reduced FAD and FMN

Glucose

2 Pyruvate 2 Lactate2 NADH

2 NAD+

Re-cycling of cytosolic NAD

Transfer of reducing equivalents via NADP cycle

NADP+ NADPH

Reductivebiosynthetic

reactions

Catabolism

Reductivebiosynthetic

product

Oxidizedprecursor

Reducedfuel

Oxidizedproduct

Glucose 6-phosphate

6-Phosphogluconate

Ribulose 5-phosphate

Ribose 5-phosphate

Nucleotides, coenzymes,DNA, RNA

Fatty acids,sterols, etc

NADPH

NADPH

NADP+

NADP+

CO2

GSSG

2 GSH

Precursors

reductivebiosynthesis

Nonoxidativephase

Oxidative phase

glutathionereductase

transketolase,transaldolase

General scheme of the pentose phosphate pathway

OH OHOH

OH

CH2OPO

O-

-O

OHOH

OH

CH2OPO

O-

-O

O

glucose 6-phosphate

glucose 6-phosphatedehydrogenase

6-phospho-glucono-δ-lactone

NADP+ NADPH + H+

Mg2+

Acetyl-CoA

CO2

NADH,FADH2

(reduced e- carriers)

ATPADP + Pi

Respiratory(electron transfer)

chain

Citricacid cycle

Glycolysis

Stage 3Electron transfer

and oxidativephosphorylation

Stage 2Acetyl-CoAoxidation

Stage 1Acetyl-CoAproduction

CO2

e-

e-

e-

e-

e-

e-

e- e-

e-

Aminoacids

Fattyacids Glucose

2H+ + 1/2O2

H2O

CO2

pyruvatedehydrogenasecomplex

OxaloacetateCitrate

Three stages of cellular respiration

Phospholipids

Triacylglycerols

Starch

Pyruvate

Sucrose

Isoleucine

LeucineSerine

Phenyl-alanine

Bileacids

Steroidhormones

Caratenoidpigments

Rubber

Isopentyl-pyriphosphate

Cholesterylesters

Vitamin K

Cholesterol

Triacylglycerols

Eicosanoids

Mevalonate

Fatty acids

CDP-diacylglycerol

Diverging anabolism

Converging catabolism

CO2

OxaloacetateCitrate

CO2

Cyclic pathway

Acetate(acetyl-CoA)

Alanine

GlucoseGlycogen

Fatty acids

Actoacetyl-CoA

Phospholipids

Types of nonlinear metabolic pathways

A

BB B

BB

B B

Enzyme-limitedreaction (far from

equilibrium)

C

Substrate-limitedreaction (at ornear equilibrium)

D

EEE

EE

EE

E

E

EEE E

EE

F G

H

H

HH

HH

H

I J

Enzyme-limited and substrate-limited reactions

enzyme 1

IsoleucineThreonineC D E FA B

Feedback inhibition

Fructose 6- + ATPphosphate

Fructose 1,6- + ADPphosphate

ATP AMP, ADP

citrate fructose 2,6-bisphosphate

Regulation of phosphofructokinase-1

Relative Changes in [ATP] and [AMP] When ATP Is Consumed

Adenine nucleotide

Concentration before ATP

depletion (mM)

Concentration after ATP depletion (mM)

Relative change

ATP 5.0 4.5 10%ADP 1.0 1.0 0AMP 0.1 0.6 600%

[Fructose 6-phosphate] (mM)

PFK

-1 a

ctiv

ity (%

of V

max

)

-F2,6BP

+F2,6BP

100

80

60

40

20

00 0.05 0.1 0.2 0.4 0.7 1.0 2.0 4.0

Role of fructose 2,6-bisphosphate in regulation of PFK-1

Glyceraldehyde3-phosphate

Fructose 1,6-bisphosphate

Dihydroxyacetonephosphate

Glycolysispathway

Gluconeogenesispathway

Fructose 6-phosphate

2-Phosphoglycerate

Phosphoenolpyruvate

3-PhosphoglycerateATP ATP

ADP ADP

NADH

NAD+

NADH

NAD+

1,3-Bisphosphoglycerate

Thirdbypass

Secondbypass

ATP

ADP

Pi

Glucose 6-phosphate

ATP

ADP

Pi

Glycogen

GlucoseGlucose 1-phosphate

UTP

UDP

AMP,fructose 2,6-

bisphosphate

fructose 2,6-bisphosphate

Glycolysis and gluconeogenesis

Compartmentalization of glycolysis, the citric acid cycle, and oxidative phosphorylation

ATP

ATP

ATP

P

NADHNADH

NADHNAD+

ADP

H2O

O2 CO2

NADH

Acetyl-CoA

Glucose

Citricacidcycle

Citric acidcycle andoxidativephosphoryla-tion in the mitochondria

Glycolysisin the cytosol

Glucose

Pyruvate

ATP

ATP

ATP

ATP

+

Vitamins and Coenzymes

Vitamin Coenzyme Form

Water-SolubleNiacin (nicotinic acid)Riboflavin (vitamin B2)Thiamine (vitamin B1)Pantothenic acidPyridoxal, piridoxine (vitamin B6)Cobalamine (vitamin B12)BiotinLipoic acidFolic acid

NAD+, NADP+

FAD, FMNThiamine pyrophosphateCoenzyme APyridoxal phosphate5’-DeoxyadenosylcobalomineBiotin-lysineLipoyl-lysineTetrahydrofolate

Fat-SolubleRetinol (vitamin A)Cholecalciferol (vitamin D3)α-Tocopherol (vitamin E)Vitamin K

N

N

NH2

CH2

CH3

SN

C

CH3

CH2 CH2 O P O P O-OO

O- O-

H+

N

N

NH2

CH2

CH3

SN

C

CH3

CH2 CH2 OH

H+

ATP+

AMP

TPP-synthetase

Thiamine (vitamin B1) Thiamine pyrophosphate (TPP)

Acidic proton

Thiamine pyrophosphate

thiazoliumring

activealdehyde

Hydroxyethyl thiamine pyrophosphate

pyruvate CO2

pyruvatedehydrogenase

N

N

NH2

CH2

CH3

SN

C

CH3

CH2 CH2 O P O P O-OO

O- O-

H

N

N

NH2

CH2

CH3

SN

C

CH3

CH2 CH2 O P O P O-OO

O- O-

C

H

OHCH3

Thiamine pyrophosphate and its role in decarboxylation

The structure of coenzyme A

Pantothenic acidβ-Mercaptoethylamine

3’-Phosphoadenosine diphosphate(3’-P-ADP)

Coenzyme A

O

OHO

CH2

N

N

N

N

NH2

P O-

O-O

OPOPOCH2CCCNCH2CH2CNCH2CH2HSH

O

H

O

H CH3

CH3OH

O- O-

O O

COO-

C

CH2

CH2

COO-

O COO-

C HH2N

R

COO-

C

CH2

CH2

COO-

HH2N COO-

C

R

O

α-Keto acidL-Amino acid L-Glutamateα-Ketoglutarate

aminotransferase+ +

PLP

Enzyme-catalyzed transaminations

NH

H2C

C

HO CH3

O

P

O-

-O O

H

O

Pyridoxal phosphate(PLP)

+NH

H2C

C

HO CH3

O

P

O-

-O O

H3N

H

H

Pyridoxaminephosphate

+ +

Pyridoxal phosphate, the prosthetic group of aminotransferases

NH

H2C

C

HO CH3

O

P

O-

-O O

N

HH

NH

H2C

C

HO CH3

O

P

O-

-O O

H

OLys NH2

LysEnz

Enz

Schiff base

+

+

+

H2O

..

Pyridoxal phosphate is bound to the enzyme through a Schiff-base linkage

HN NH

S(CH2)4 C

N

O

O

H

Enz

Enz

HN NH

S(CH2)4 C

NH

O

O

Enz

-O P O P O P O

O- O- O-

O O O

AdenineRib

Bicarbonate

ATP

Pyruvate enolate

Carboxybiotinyl-enzyme

Biotinyl-enzyme

Biotinyl-enzyme

Oxaloacetate

+

ADP + Pi

1

2

-O CO

OH

H

C

O

-O N NH

S(CH2)4 C

N

O

O

C C-O

O O-

CH2C C-O

O O

CH2-

CO

OHC C

-O

O O

CH2

Role of biotin in carboxylation reactions

CHCH2

CH2

CH2

S

HS

CCH3

O

Polypeptide chain ofE2 (dihydrolipoyltransacetylase)

Lipoicacid

Acetylatedform

Reducedform

Oxidizedform

Lysresidue

of E2

CHCH2

CH2

CH2

HS

HS

S

S CHCH2

CH2

CH2

CH2

CH2

CH2

C O

HNCH2

CH2

CH2

CH2

CHNH C

O

Lipoic acid (lipoate) in amide linkage with Lys residue

Tetrahydrofolate (H4 folate)6-methylpteringlutamate

p-aminobenzoate1

23

4 5 6

78

9

8a

104aN

N N

N

H2N

O

H

CH2

H

H

NH C

O

NH CH

COO-

CH2 CH2 COO-H

H

H

Tetrahydrofolate (H4 folate)

COO-

C HH2N

H

NADH

NADH

+ H+

NAD+

NAD+

PLP

Serine Glycine

TetrahydrofolateN5,N10-Methylenetetrahydrofolate

N5,N10-Methyltetrahydrofolate

N5-Formyletrahydrofolate

(most reduced)

(most oxidized)

Oxidation state(group transferred)

N5,N10-Methylene-tetrahydrofolate

reductase

N5,N10-Methenyl-tetrahydrofolate

reductase

+ H+

H2O

serine hydroxymethyl transferaseN

CH2N

CH2

H

NH

H

H

COO-

C HH2NCH2OH

N

CH2N

CH2

H

NC

H

H2

N

CH2N

CH2

H

NCH3

H

H

CH3

CH2OH

N

CH2N

CH2

H

NCO H

H

H

CO

H

Conversion of one-carbon units on tetrahydrofolate

O

OH O

CH2N

N N

N

NH2

H

N

N

N

N

CH2

CH2NO

CH3

CH2

CH2

CO

NH2

H CH2

CH3

C

O

NH2

CH2

H

H CO

NH2

CH3

H2CH2C

C

H3C

CH3

O

H2N

H

H2CH2C

CO

NH2

CH3CH3 Co3+

CH2

CH2

CNHCH2

HC CH3

OP O-

OO

N

NCH3

CH3

O

CH2OHOH

O

5'-Deoxy-adenosine

Amino-isopropanol

Corrinringsystem

Dimethyl-benzimidazoleribonucleotide

1’

2’ 3’

4’

5’

Coenzyme B12 is the cofactor form of vitamin B12

-O P O P O P O-O O O

O- O-O-

Co

Coenzyme B12

Cobalamin

1'

2' 3'

4'

5'

ATP

O

OH O

CH2 O P O P O P O-O O O

O- O-O-N

N N

N

NH2

Co

O

OH O

CH2N

N N

N

NH2

The formation of coenzyme B12 occurs in reaction in which triphosphate is cleaved from ATP

Coenzyme B12-catalyzed group transfer

methylmalonyl-CoAmutase

Coenzyme B12 C

H

H C

H

CO-

O

HCO S-CoA

C

H

H C

H

H

CO

CO-

O

S-CoA

L-Methylmalonyl-CoA Succinyl-CoA

Coenzyme B12C C

H X

CC

HX

all-trans-Retinoic acid

C

CH3

CH3

CH3

CH3

CH3

OHO

β-Carotene

CH3

CH3CH3

CH3

CH3

CH3

CH3

CH3

CH3

CH3

point ofcleavage

oxidation ofalcohol toaldehyde

oxidationof aldehyde

to acid

visiblelight Vision

Hormonalsignal

Vitamin A1(retinol)

CH2OH

CH3

CH3

CH3

CH3

CH3

26

7

11

15

cleavage

isomerization/oxidation ofalcohol toaldehyde

11-cis-Retinal(visual pigment)

CH3

CH3

CH3

CH3

C

H3C

O H

11

12

all-trans-Retinal

11

12

C

CH3

CH3

CH3

CH3

CH3

OH

all-trans-Retinal

11

12

C

CH3

CH3

CH3

CH3

CH3

OH

Vitamin A1 and its precursors and derivatives

7-Dehydrocholesterol

UV light

2 steps in skin

Cholecalciferol (vitamin D3)

1 step in the liver

1 step in the kidney

1,25-Dihydroxycholecalciferol (1,25-dihydroxyvitamin D3)

H3C CH3

CH3CH3

CH3

HO

12

34

56

7

8

9

10

H3CCH3

CH3CH3

HO

CH2

12

3

45

67

H3CCH3

CH3CH3

HO

CH2

OH

OH

25

12

3

45

67

Vitamin D3 production and metabolism

O

CH

CH2 C CH3

O

OH

Warfarin: a bloodanticoagulant

Vitamin E: an antioxidantO

CH2

CH3

CH2 CH2 CH CH2

CH3

CH2 CH2 CH

CH3

CH2 CH2 CH2 CH

CH3

CH3

CH3

CH3

CH3

HO

Vitamin K1: a blood-clottingfactor (phylloquinone) CH2 CH2 CH

CH3

CH3CH2 C CH2

CH3

(CH2 CH2 CH

CH3

CH2)2CH2

O

O

CH3

Ubiquinone: a mitochondrialElectron carrier (coenzyme Q)

CH2 C CH3

CH3

CH2CH2 C CH2)n

CH3

(CH2

CH3

CH2 C CH2

CH3

CH2CH3O

CH3O

O

O

Vitamins E and K and the lipid quinones are oxidation-reduction cofactors

Vitamin K stimulates the carboxylation of glutamate residues

N CH C

H CH2

H2C C O-

O

O

N CH C

H CH2

C C O-

O

O

C-O

O

KH2 KO

K

Glu γ-carboxy-Glu

Warfarin

warfarinsensitive

O

CH

CH2 C CH3

OOH

γ-glutamylcarboxylase

vitamin K epoxide reductase

vitamin K reductase

12

2

1

1. Living cells constantly perform work. They require energy for maintaining their highly organized structures, synthesizing cellular components, generating electric currents, and many other processes

2. ATP is a chemical link between catabolism and anabolism. The exorgonic conversion of ATP to ADP and Pi, or to AMP and PPi is coupled to many endergonic reactions and processes. ATP provides the energy for anabolic processes through the group transfer reactions. In many organisms, a central energy-conserving process is the stepwise oxidation of glucose, fatty acids, and amino acids to CO2, in which some of the energy of oxidation is conserved in ATP as electrons are passed to oxygen

3. Many biological oxidation reactions are dehydrogenations in which one or two hydrogen atoms are transferred from a substrate to a hydrogen acceptor

4. NAD and NADP are the freely diffusible coenzymes of many dehydrogenases. They accept two electrons and one proton. FAD and FMN, the flavin nucleotides, serve as tightly-bound prosthetic groups of flavoproteins. They can accept either one or two electrons and one or two protons

5. Vitamins are essential nutrients that are required in the diet because they cannot be synthesized by the organism itself. Often they are components or precursors of coenzymes