BIOENERGETICS, ENZYMES, & METABOLISM

Chapter 3

What is Bioenergetics?
The study of energy transformations occurring in

living organisms

 Energy Capacity to do work, or to change or

move something
Thermodynamics the study of the changes in

energy that accompany events in the universe

Laws of Thermodynamics

1st Law of Thermodynamics

Energy cannot be created or destroyed only

transferred from one form to another

Total amount of energy in the universe is

constant
Examples of Energy Transduction Chemical mechanical (moving organelles) Chemical electrical (proton gradients) Chemical thermal (muscle contraction) Chemical light (fireflies & fish)

The System vs. The Surroundings

The universe can be divided into systems &

surroundings
System a subset or section of the universe

under study
Surroundings everything else The energy of the system internal energy (E)
Its change during a transformation is called E

Energy Conversion & Heat

When there is energy conversion (E) in a system,

heat content (Q) may increase or decrease

Reactions that lose heat are exothermic

Reactions that gain heat are endothermic

The energy conversion in a system can be

calculated with a mathematical formula

E = Q W

E Can Be Positive or Negative

The 2nd Law of Thermodynamics

Events in the universe tend to proceed from a state of

higher energy to a state of lower energy

Such events are called spontaneous, they can occur

without the input of external energy

Exergonic Release or Produce energy during reaction Endergonic Uses or Requires energy for reaction to proceed

Entropy (S) Measure of randomness & disorder It is the energy NOT available to do work

Entropy in the Universe is Increasing

Entropy increases with

every energy transfer

Entropy is associated

with random movements of particles or matter

Living systems maintain

a state of order, or low entropy

But Living Organisms Are OrganizedHow Do You Explain That?

During every energy transfer (even nonspontaneous ones) heat is lost to the

environment
Heat = energy in most disordered form

Therefore the total entropy (Cell + environment) increases

Free Energy (G)

The 1st and 2nd laws of thermodynamics can be

combined and expressed mathematically

Equation: H = G + TS

Total energy (H) = Free Energy (G) + Entropy (TS)

Spontaneity
If G < 0 then the rxn is favorable If G > 0 then the rxn is not favorable

Free-Energy Changes in Chemical Rxns

All chemical reactions are theoretically reversible All chemical reactions spontaneously proceed

toward equilibrium (Keq = [C][D]/[A][B])

The rates of chemical reactions are proportional

to the concentration of reactants

At equilibrium, the free energies of the products

and reactants are equal (G = 0)

 Standard free energy changes are calculated based on

equilibrium conditions
G = -RT ln Keq G = -RT ln [C] [D]/[A][B]
Standard conditions are not representative of cellular conditions,

but are useful to make comparisons

Non-standard conditions are corrected for

prevailing conditions
G = G + RT ln Keq G = G + RT ln [C] [D]/[A][B]

Conditions in the Cell are NOT Standard

Laws of thermodynamics say rxns are

spontaneous/favorable
Unfavorable reactions are still necessary Unfavorable reactions occur b/c of coupled reactions

Coupling Endergonic & Exergonic Rxns

Example of Endergonic & Exergonic Rxns

Glutamic acid + NH3 Glutamate Endergonic G = +3.4 kcal/mol unfavorable

ATP + H2O ADP + Pi Exergonic G = -7.3 kcal/mol

Glutamic acid + NH3 + ATP Glutamate + ADP + Pi G = -3.9 kcal/mol

Equilibrium vs. Steady-State Metabolism

Cellular metabolism is

nonequilibrium metabolism
Cells are open

thermodynamic systems
Cellular metabolism

exists in a steady state

ENZYMES
Biological Catalysts

Basic Properties of Enzymes

Required only in small amounts Are not permanently altered during the course of a reaction Do NOT affect the thermodynamics of rxns, only the rates Are highly specific for their substrates

Produce only appropriate metabolic products

Can be regulated to meet the needs of a cell

Why Dont Thermodynamically Favorable Reactions Happen on Their Own?

Molecules are in a relatively stable state Bonds are not going to break or form

without the input of some energy

There is an energy barrier that prevents

the reaction from proceeding

Activation Energy (EA) The amount of kinetic energy needed to start a rxn

Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

Energy State of Reaction

Reactant Initial state

Energy of activation in the absence of enzyme Energy of activation in the presence of enzyme

Products Final state

Progress of Reaction

Enzymes Lower the Activation Energy

No enzyme = only a

few substrate converted (Pink/Orange)

Enzyme = large

proportion of substrate converted (Blue)

The Enzymatic Reaction

Substrate binds to a portion of the enzyme active site

 Substrate & active

sites have complementary shapes

Substrate is often

taken into a hydrophobic cleft of the enzyme

Mechanisms of Enzyme Catalysis

1. Substrate Orientation Optimal position for rxn

2. Changing Substrate Reactivity R groups manipulate interactions

3. Inducing Strain in the Substrate Stresses covalent bonds

Inducing Strain in the Substrate

Shifts in the

conformation after binding cause an induced fit between enzyme and the substrate
Covalent bonds of the

substrate are strained

Destabilizes substrate
Substrate must adopt

transition state

Some Common Types of Enzymes

Enzyme Hydrolases Nucleases Proteases Synthases Isomerases Polymerase Kinases Phosphatases Oxido-Reductases ATPases Reaction Catalyzed Catalyze a hydrolytic cleavage reaction Break down nucleic acid by hydrolyzing nucleotides Break down proteins by hydrolyzing amino acids Synthesize molecules in anabolic reaction sby condensing two smaller molecules together Catalyze the rearrangement of bonds within a single molecule Catalyze polymerization reaction such as the synthesis of DNA Catalyze the addition of phosphate groups to molecules Catalyze the hydrolytic removal of a phosphate group from a molecule Catalyze reactions in which one molecule is oxidized while the other is reduced Hydrolyze ATP

Enzymes May Have Helpers

May be conjugated with

nonprotein components
Cofactors
Inorganic metals that help bring the

active site & substrate closer together

Coenzymes
Organic compounds that help

perform a chemical alteration to the substrate

Some Common Cofactors

Some Common Coenzymes

Vitamin Thiamine (Vitamin B1) Riboflavin (Vitamin B2) Niacin Coenzyme Thiamine pyrophosphate FADH NADH, NADPH Enzyme-Catalyzed Reaction Requiring These Coenzymes Activation & transfer of aldehydes Oxidation-reduction Oxidation-reduction

Pantothenic acid
Pyridoxine Biotin

Coenzyme A
Pyridoxal phosphate Biotin

Acyl group activation & transfer

Reactions involving amino acid activation CO2 activation & transfer

Lipoic acid
Folic acid Vitamin B12

Lipoamide
Tetrahydrofolate

Acyl group activation; oxidationreduction

Acitvation & transfer of single carbon groups

Cobalamin coenzymes Isomerization & methyl group transfers

Enzyme Kinetics
The rate at which enzymes catalyze reactions Must be determined under controlled conditions Enzyme kinetics is interested in the initial velocitythe speed at

which the enzyme works before any product is produced

Rate of enzymatic reaction will increase until the enzyme has

reached saturation
Saturation = enzymes maximum capacity This statement assumes that substrate conc. is increasing

 Vmax = Speed of rxn at saturation

Turnover # = max # of molecules that can be converted to product by 1

enzyme
The Michaelis constant (KM) can tell the affinity of enzyme for the substrate
(KM) = substrate conc. at of Vmax

Lineweaver-Burk Plot can make it easier to determine Vmax and KM.

Temperature & pH can Affect Enzymatic Reaction Rates

Enzyme Inhibitors
Inhibitors slow down enzymatic rxns
Irreversible inhibitors
Tightly bound to the enzyme

Reversible inhibitors
Loosely bound to the enzyme

Competitive vs. Noncompetitive Inhibition

Competitive
Similar in structure to

Noncompetitive
Bind to sites other than

substrate
Bind to the active site to

active sites and inactivate the enzyme

The maximum velocity of

slow enzyme activity

Can be overcome with

enzyme molecules cannot be reached

Cannot be overcome with

high substrate/inhibitor ratios

high substrate/inhibitor ratios.

METABOLISM

Metabolism Overview
Chemical rxns are organized into metabolic

pathways
Anabolism Assembling/building macromolecules that the cell needs
Uses/consumes energy

Catabolism Breaking down/destroying organic compounds to obtain energy

Releases/makes energy

Metabolic Pathways

Metabolic rxns occur in a series of steps The products of some rxns are the reactants/substrates for the next Each step is catalyzed by an enzyme Enzymes of a metabolic pathway are usually confined to one location in a cell

Anabolic & Catabolic Pathways are Interconnected

The Capture & Utilization of Energy

Cells obtain energy by oxidizing organic molecules

Exergonic reaction a rxn that produces or

releases energy (G < 0)

Endergonic reaction a rxn that requires or uses

energy (G > 0)

Transferring Electrons Moves Energy

Oxidation Reduction Reactions

OIL RIG
Redox reactions always occur in pairs An electron donor = reducing agent (red) An electron acceptor = oxidizing agent (blue)

eReduced Oxidized Oxidized

eReduced

e-

e-

Fe0

Cu2+

Fe2+

Cu0

Activated Carriers
The energy released from the oxidation of organic

molecules must be temporarily stored

Energy is in an easily exchangeable formlike $$$ ATP, NADH, FADH2, NAPDH Diffuse rapidly in the cell Carry energy from one site to another
e- e-

CELLS USE ENERGY IN FOOD TO MAKE ATP

Glycolysis

Glycolysis
Probably the most ancient metabolic pathway Splitting of Sugar

ATP Glucose (6C) 1 2 3 4 5 6

Uses & makes

2 Phases Energy Investment Energy Production No O2 needed

Pyruvate (3C)

Pyruvate (3C)

Glycolysis and ATP Formation

10 Rxns All but 3 are near

equilibrium (G ~ 0) under cellular conditions

These 3 rxns drive

glycolysis

 Transfer Potential
ATP formation is only

moderately endergonic compared with other phosphate transfer in cells

Transfer potential shown when

molecules higher on the scale have less affinity for the group being transferred than are the ones lower on the scale.
The less the affinity, the better

the donor.

Glycolysis and ATP Formation Cont.

Glucose is phosphorylated to glucose 6-phosphate by

using ATP
Glucose 6-phosphate is isomerized to fructose 6-

phosphate
Fructose 6-phosphate is phosphorylated to fructose 1,6-

bisphophate using another ATP

Fructose 1,6-bisphosphate is split into two three-carbon

phosphorylated compounds.

Glycolysis and ATP Formation Cont.

NAD+ is reduced to NADH when glyceraldehyde 3-

phosphate is converted to 1,3-bisphosphoglycerate

Dehydrogenase enzymes oxidize and reduce cofactors

NAD+ is a nonprotein cofactor associated with

gluceraldehyde phosphate dehydrogenase

NAD+ can undergo oxidation and reduction at different

places in the cell

NADH donates electrons to the electron transport chain in

the mitochondria

Glycolysis and ATP Formation Cont.

ATP is formed when 1,3-bisphosphoglycerate is

converted to 3-phosphoglycerate by 3-phosphoglycerate kinase

Kinase enzymes transfer phosphate groups Substrate-level phosphorylation occurs when ATP is formed by a

kinase enzyme

3-phosphoglycerate is converted to pyruvate via three

sequential reactions, in one of them a kinase phosphorylates ADP

Glycolysis and ATP Formation Cont.

Glycolysis can generate a net of 2 ATPs for each glucose
Glycolysis occurs in the absence of oxygen, it is an

anaerobic pathway
The end product, pyruvate, can enter aerobic or

anaerobic catabolic pathways.

Pyruvate is Versatile

Anaerobic Oxidation of Pyruvate: The Process of Fermentation

Fermentation restores NAD+ from NADH
Under anaerobic conditions, glycolysis depletes the

supply of NAD+ by reducing it to NADH pyruvate

In fermentation, NADH is oxidized to NAD+ by reducing

In muscle and tumor cells pyruvate is reduced to lactate

In yeast and other microbes, pyruvate is reduced and

converted to ethanol

Fermentation is inefficient with only about 8% of the

energy of glucose captured as ATP

The Process of Fermentation

Reducing Power
Anabolic pathways require a source of electrons to form

larger molecules
NADPH donates electrons to form large biomolecules
NADPH is a coenzyme similar to NADH
The supply of NADPH represents the cells reducing power NADP+ is formed by phosphate transfer from ATP to NAD+

Reducing Power Cont.

NADPH and NADH are interconvertible, but have

different metabolic roles

NADPH is oxidized in anabolic pathways NAD+ is reduced in catabolic pathways

The enzyme transhydrogenase catalyzes the transfer

of hydrogen atoms from one cofactor to the other

NADPH is favored when energy is abundant NADH is used to make ATP when energy is scarce

Metabolic Regulation
Cellular activity is regulated as needed Regulation may involve controlling key enzymes of metabolic

pathways
Enzymes are controlled by alteration in active sites
Covalent modification of enzymes regulated by phosphorylation

such as protein kinases

Allosteric modulation by enzymes regulated by compounds

binding to allosteric sites.

In feedback inhibition, the product of the pathway allosterically

inhibits one of the first enzymes of the pathway.

Feedback Inhibition
Build up of product has a negative effect Turns pathway off

Separating Catabolic & Anabolic Pathways

Glycolysis and gluconeogenesis are the catabolic

and anabolic pathways of glucose metabolism

Synthesis of fructose 1,6-bisphosphate is coupled to

hydrolysis of ATP

Breakdown of fructose 1,6-bisphosphate is via hydrolysis by

fructose 1,6-bisphosphatase in gluconeogenesis ATP as the allosteric inhibitor

Phosphofructokinase is regulated by feedback inhibition with Fructose 1,6-bisphosphatase is regulated by covalent

modification using phosphate binding

ATP levels are highly regulated

Separating Catabolic & Anabolic Pathways

Separating Catabolic and Anabolic Pathways

Anabolic pathways do not proceed via the same reactions

as the catabolic pathways even though they may have steps in common
Some catabolic pathways are essentially irreversible due to large

G values
Irreversible steps in catabolic pathways are catalyzed by different

enzymes from those in anabolic pathways