BIOCHEMISTRY

František Vácha

http://www.prf.jcu.cz/~vacha/

JKU, Linz

Recommended reading:

D.L. Nelson, M.M. Cox

Lehninger Principles of Biochemistry

D.J. Voet, J.G. Voet, C.W. Pratt

Principles of Biochemistry

L. Stryer

Biochemistry

April

4. 4.

11. 4.

18. 4.

25. 4.

May

2. 5.

9. 5.

23. 5.

30. 5.

June

6. 6.

13. 6.

20. 6.

27. 6.

March

7. 3.

14. 3.

1. What are the chemical and three-dimensional structures of biological molecules

2. How do biological molecules interact with each other

3. How does the cell synthesize and degrade biological molecules

4. How is the energy conserved and used by the cell

5. What are the mechanisms for organizing biological molecules and coordinating

their activities

6. How is genetic information stored, transmitted and expressed

Principle issues of Biochemistry

Biochemistry reveals the working mechanisms of the natural world

Life and Cells

B, F, Al, Si, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Br, Mo, Cd, I, W

Simple inorganic

compounds form

more complex

molecules, that are

the basic of live

forms

• in molecules

• as ions

Biogenic elements

Living organisms are

based on various complex

molecules consisting of

simple atoms

Combining different functional groups

in a single large molecule increases the

chemical versatility of such molecule

Different macromoleculs with

complementary arrangements of

functional groups can associate with

even greater range of functional

possibilities

Thermodynamics

and

Spontaneity of biochemical reactions

Gibbs free energy

ΔG = ΔH – TΔS

ΔG = ΔGo + RT lnK

(ΔGo = – RT lnKeq)

ΔH – Enthalpy - heat at constant pressure (exothermic, endothermic)

T – temperature in Kelvins

S – Entropy

R – gas constant

K – reaction quotient

Keq – equilibrium constatnt

Go – Standard free energy

Spontaneity of biochemical reactions

Equilibrium constant measures the

direction of spontaneous processes

At biochemical standard conditions (1M, pH 7, 298 K, 101.3 kPa)

the free-energy change of a biochemical reaction is simply an

alternative expression of the equilibrium constant

Actual free-energy changes depend

on reactant and product

concentrations

Standard equilibrium (K’eq) – initial concentrations of each

component is at 1M

This is not the case of living organism

Different concentrations of metabolites can affect the

reaction direction

In human erythrocytes

ATP = ADP + Pi

ATP = 2.25 mM

ADP = 0.25 mM

Pi = 1.65 mM

T = 37 oC (310 K)

DG’o = - 30.5 kJ/mol

DG = - 52 kJ/mol

Adenosin nucleotide and inorganic

phosphate concentrations in some cells

• Large negative value of ΔG does not ensure that a

process will proceed at measurable rate

• The rate depends on the detailed mechanism of the

reaction and not on the ΔG

• Nearly all molecular components of an organism can

react with each other and many of these reactions are

thermodynamically favored

• Organism can regulate the reactions by altering their

mechanisms

• Enzyme catalysis

Life Needs Energy

• The ultimate source of this energy on the Earth is

the sunlight

Organisms can be classified according to the

source of energy and carbon

Water

and

noncovalent weak forces

~ 70 % of human body mass is water

• medium for majority of biochemical reactions

• water itself actively participates in many biochemical

reactions

• nearly all biological molecules acquire their shape,

and therefore their functional properties, in an

interaction with water

• the unique physical and chemical properties of

water enables the present life forms on the Earth

Water

Hydrogen bonds – key feature of water for biology

Water is polar molecule: - 0.66 e on oxygen and + 0.33 e on each hydrogen

Hydrogen bond in water is ~ 1.9 Å

Energy of H-bond ~ 20 kJ . mol-1

—F—H…..:F— 155 kJ/mol 1.13 Å

—O—H…..:N— 29 kJ/mol 2.88 Å

—O—H…..:O— 21 kJ/mol 2.70 Å

—N—H…..:N— 13 kJ/mol 2.93 Å

—N—H…..:O— 8 kJ/mol 3.04 Å

Noncovalent - weak forces are the principal

interactions in biological molecules

the whole life is based on weak interactions

• Biogenic elements are part of complex molecules or

appear as ions

• Chemical versatility of macromolecules with different

functional groups

• Gibbs free energy as a measure or reaction spontaneity

• Water as a basic environment for biochemical reactions

• Noncovalent – weak forces are the key interactions in

biomolecules

• H-bonds

Learning objectives

Introduction to metabolism

Metabolism

• Sum of all chemical reactions in an

organism

• Complex and highly coordinated

• The core parts are similar in all living

organisms

• Reactions in sequence form

metabolic pathways

• Some pathways are primarily

targeted to produce energy

• catabolism

• Some pathways are primarily

targeted to synthetize new

substances (on the cost of energy)

• anabolism

Catabolism of proteins, fats,

and carbohydrates in the

three stages of cellular

respiration

• Stage 1: oxidation of fatty acids, glucose, and

some amino acids yields acetyl-CoA.

• Stage 2: oxidation of acetyl groups in the citric

acid cycle to form NADH and FADH2

• Stage 3: electrons are funneled into a chain of

electron carriers reducing O2 to H2O. This

electron flow drives the production of ATP.

Complete Oxidation of Reduced

Compounds is Strongly Favorable

• This is how chemotrophs obtain most of their energy

• In biochemistry the oxidation of reduced fuels with O2 is stepwise and controlled

• Thermodynamically favorable is not the same as being kinetically rapid – enzyme catalysis

Electron carriers

• A few types of coenzymes and proteins

serve as universal electron carriers

• Many biochemical oxidation-reduction

reactions involve transfer of two electrons

• In order to keep charges in balance, proton

transfer often accompanies electron

transfer

NAD and NADP as common

redox cofactors

• These are commonly called pyridine nucleotides

• They can dissociate from the enzyme after the reaction

• In a typical biological oxidation reaction, hydride(:H-) from an alcohol is transferred to NAD+ giving NADH

• AH2 + NAD(P)+ A + NAD(P)H + H+

NAD and NADP in metabolism

NAD+/NADH - catabolism, further in ATP

production

NADP+/NADPH – anabolism, biosynthetic

reactions

Flavin Cofactors allow Single

Electron Transfers

• Flavoproteins (FMN, FAD)

• May participate in one- or two-electron transfers

• Flavin cofactors are usually tightly bound to proteins, some covalently

• Variability in reduction potentials

Iron-Sulfur Centres

• Bound in proteins

• Transfer one electron i time

• Diferent types

Cytochromes

• Membrane or soluble heme-containing protein

• Heme – a tetrapyrrol binding an iron ion in a form

of either ferrous (Fe3+, oxidized) or ferric(Fe2+,

reduced)

• Single electron carriers

Principal role of ATP

in metabolism

• stores energy obtained in catabolic reactions

• transport the energy to compartments or parts of organism where it is needed

• provides the energy for anabolic biosynthetic processes

Chemical basis of large negative

free-energy of ATP

• Separation of negative charges on phosphate

oxygens upon ATP hydrolysis

• Resonance stabilization of phosphate products

• Ionisation of ADP product

• Better solvation of products

ATP provides energy by group transfer

Simple hydrolysis of ATP is not the

source of energy (only liberation of

heat)

• In most cases it is two-step

process:

1) Favorable ATP hydrolysis and Pi

transfer

2) Resonance stabilization of free Pi

• Some processes involve simple

hydrolysis:

- Binding ATP to a protein and its

hydrolysis – conformation change

of the protein – mechanical motion

Actual DG of ATP hydrolysis

depends on a type of tissue

The cellular concentration of ATP is usually above the equilibrium

constant making it even better source of energy

Actual DG of ATP hydrolysis

depends on a type of tissue

DG = -30.5 kJ/mol + [(8.315 kJ/mol.K)(310 K) ln((0.25x10-3)(1.65x10-3))/(2.25x10-3)

DG = -52 kJ/mol

Mg2+ binds to ATP and ADP to form complexes of Mg-ATP

and Mg-ADP

Regulatory role, shielding of negative charges of oxygen,

conformation changes of ATP and ADP molecules

The Role of Magnesium in ATP Reactions

Several Phosphorylated Compounds

Have Larger DG’° Than ATP

• Again, electrostatic repulsion within the reactant, molecule is relieved

• The products are stabilized via resonance, or by more favorable solvation

• Possible tautomerization product

Hydrolysis of phosphoenolpyruvate (PEP)

Hydrolysis of 1,3 bisphosphoglycerate

Hydrolysis of phosphocreatine

Substrate level phosphorylation

Phosphorylated molecules with higher ΔG°’ can be used to synthesize ATP

PEP + ADP = Pyruvate + ATP

ΔG°’ – 61,9 kJ/mol

Hydrolysis of Thioesters

• Acetyl-CoA

Hydrolysis of Thioesters

• Hydrolysis of thioesters, such as acetyl-CoA is

strongly favorable

• Acetyl-CoA is an important donor of acyl groups

– Feeding two-carbon units into metabolic pathways

– Synthesis of fatty acids

Hydrolysis of acetyl-coenzyme A

• Catabolism and Anabolism

• Stepwise oxidation as a source of energy

• Electron carriers in biological systems

• Principal role of ATP in metabolism

• Energy in ATP

• Substrate level phosphorylation

Learning objectives