Enzymes

Bio 11, November 9, 2007Next Tuesday- last day to withdraw

Which is the most oxidized?

Methane Methanol Formaldehyde Carbon dioxide

Which is the most reduced?Which is highest in energy?Which is lowest in energy?

Glycolysis is actually 10 chemical reactions

Each step in the reaction is catalyzed by a different enzyme

1 Glucose molecule yields 2 ATP, 2 NADH, 2 Pyruvate

Pyruvate enters the mitochondrion, loses a carbon, and binds to coenzyme A to enter Krebs cycle

CYTOSOL

Pyruvate

NAD+

MITOCHONDRION

Transport protein

NADH + H+

Coenzyme ACO2

Acetyl Co A

•The Krebs cycle is a set of many reactions•Each reaction is catalyzed by an enzyme•Such systems are called a metabolic pathways

•Net result: 2ATP, 3 NADH, 1 FADH2/Glucose

The electron transport system Generates a proton gradient, which powers ATP synthase

Protein complexof electroncarriers

H+

ATP ATP ATP

GlycolysisOxidative

phosphorylation:electron transportand chemiosmosis

Citricacidcycle

H+

Q

IIII

II

FADFADH2

+ H+NADH NAD+

(carrying electronsfrom food)

Innermitochondrialmembrane

Innermitochondrialmembrane

Mitochondrialmatrix

Intermembranespace

H+

H+

Cyt c

IV

2H+ + 1/2 O2 H2O

ADP +

H+

ATP

ATPsynthase

Electron transport chainElectron transport and pumping of protons (H+),

Which create an H+ gradient across the membrane

P i

ChemiosmosisATP synthesis powered by the flow

of H+ back across the membrane

Oxidative phosphorylation

Net result: ~32-34 ATP/glucose molecule

Complete the table

Step of respiration

Input Output(s) ATP formed

Glycolysis GlucoseKrebs cycleElectron transport/ oxidative phos.

Complete the table

Step of respiration

Input Output(s) ATP formed

Glycolysis Glucose 2 pyruvate, 2 NADH

2 (net)

Krebs cycleElectron transport/ oxidative phos.

Complete the table

Step of respiration

Input Output(s) ATP formed

Glycolysis Glucose 2 pyruvate 2 (net)Krebs cycle 2 acetyl CoA 1 FADH, 3 NADHElectron transport/ oxidative phos.

Complete the table

Step of respiration

Input Output(s) ATP formed

Glycolysis Glucose 2 pyruvate, 2 NADH

2 (net)

Krebs cycle 2 acetyl CoA 2 FADH2, 8 NADH, 2 CoA, 6CO2

2

Electron transport/ oxidative phos.

2 FADH2, 10 NADH, 6O2

6H2O ~32-34

Complete the tableStep of respiration

Input Output(s) ATP formed

Glycolysis Glucose 2 pyruvate, 2 NADH

2 (net)

Krebs cycle 2 acetyl CoA 2 FADH, 6NADH, 2 CoA, 6CO2

2

Electron transport/ oxidative phos.

2 FADH, 6 NADH, 6O2

6H2O ~32-34

Total C6H12O6 + 6O2 6H2O, 6CO2 36-38

Enzymes are protein catalysts

• Catalyst- something that speeds up the rate of a reaction without being consumed by the reaction

• They are shape-dependent

• They are specific to a substrate

• Most enzyme names end with “–ase”

Diagramming a chemical reaction

• Y axis = energy in chemical bonds of products and reactants

• X axis = Reaction progress ( ≈ time)

• Diagram shows an exergonic reaction

• Ea = activation energy

Enzymes can dramatically lower the activation energy for a reaction

Reaction Course

Energyreactants

products

E a

E a

no enzyme

with enzyme

12Note that the equilibrium of the reaction is unaffected

Substrate Binding and Reaction

Example:

-galactosidase

11

lactose

galactose

glucose

H2O

- galactosidase(aka lactase in humans)

-galactosidase

10

How enzymes work

• Structure aids function

• An active site (aka “activation site”) naturally fits substrate

• Enzymes stabilize transition state of substrates

LE 8-17

Enzyme-substratecomplex

Substrates

Enzyme

Products

Substrates enter active site; enzymechanges shape so its active siteembraces the substrates (induced fit).

Substrates held inactive site by weakinteractions, such ashydrogen bonds andionic bonds.

Active site (and R groups ofits amino acids) can lower EA

and speed up a reaction by• acting as a template for substrate orientation,• stressing the substrates and stabilizing the transition state,• providing a favorable microenvironment,• participating directly in the catalytic reaction.

Substrates areconverted intoproducts.

Products arereleased.

Activesite is

availablefor two new

substratemolecules.

Ways in which Enzymes help• The active site can lower an EA barrier by– Orienting substrates correctly– Straining substrate bonds– Providing a favorable microenvironment– Covalently bonding to the substrate

ENZYMES CANNOT:-Act as an energy source-Turn an unfavorable reaction into a favorable one

Effects of Local Conditions on Enzyme Activity

• An enzyme’s activity can be affected by:– General environmental factors, such as temperature

and pH– Chemicals that specifically influence the enzyme

Effects of Temperature and pH

• Enzymes have an optimal temperature and pH

• Tertiary structure can be radically altered by changes in pH or temp

LE 8-18

Optimal temperature fortypical human enzyme

Optimal temperature forenzyme of thermophilic (heat-tolerant bacteria)

Temperature (°C)

Optimal temperature for two enzymes

0 20 40 60 80 100

Rate

of r

eacti

on

Optimal pH for pepsin(stomach enzyme)

Optimal pHfor trypsin(intestinalenzyme)

pH

Optimal pH for two enzymes

0

Rate

of r

eacti

on

1 2 3 4 5 6 7 8 9 10

Cofactors• Cofactors are

nonprotein enzyme helpers

• Coenzymes are organic cofactors

• Many vitamins are cofactors for enzymes

• Metal ions also can be essential to enzyme function

Enzyme Inhibition

• Competitive inhibitors bind to the active site of an enzyme, blocking the substrate

• Noncompetitive inhibitors (aka allosteric) away from active site, changing enzyme shape

• Many drugs are enzyme inhibitors (COX2 inhibitors, etc.)

• Some toxins bind enzymes permanently, destroying them

Regulation of enzyme activity helps control metabolism

• Chemical chaos would result if a cell’s metabolic pathways were not tightly regulated

• To regulate metabolic pathways, the cell switches on or off the genes that encode specific enzymes

Allosteric Regulation of Enzymes

• Allosteric regulation is the term used to describe cases where a protein’s function at one site is affected by binding of a regulatory molecule at another site

• Allosteric regulation may either inhibit or stimulate an enzyme’s activity

• Cooperativity is a form of allosteric regulation that can amplify enzyme activity

• In cooperativity, binding by a substrate to one active site stabilizes favorable conformational changes at all other subunits

LE 8-20b

Substrate

Binding of one substrate molecule toactive site of one subunit locks allsubunits in active conformation.

Cooperativity another type of allosteric activation

Stabilized active formInactive form

Feedback Inhibition• In feedback inhibition, the end product of a metabolic

pathway shuts down the pathway• Feedback inhibition prevents a cell from wasting

chemical resources by synthesizing more product than is needed

Active siteavailable

Initial substrate(threonine)

Threoninein active site

Enzyme 1(threoninedeaminase)

Enzyme 2

Intermediate A

Isoleucineused up bycell

Feedbackinhibition Active site of

enzyme 1 can’tbindtheoninepathway off

Isoleucinebinds toallostericsite

Enzyme 3

Intermediate B

Enzyme 4

Intermediate C

Enzyme 5

Intermediate D

End product(isoleucine)

Specific Localization of Enzymes Within the Cell

• Structures within the cell help bring order to metabolic pathways

• Some enzymes act as structural components of membranes

• Some enzymes reside in specific organelles, such as enzymes for cellular respiration being located in mitochondria

LE 8-22

Mitochondria,sites of cellular respiration

1 µm