Enzymes (Ch. 6)

• Intro

• Basics of catalysis

• General types of catalysis

• Quantification of catalysis– enzyme kinetics and inhibition

• Specific examples

• Allostery and enzyme regulation

EN

ER

GY

(G

°)

REACTION PROGRESS

G < 0

Reaction should bespontaneous

Equil should favorproducts

Biological reaction:sugar + oxygen ↔ CO2 + water

Reactants (R)

Activation energy

EA

Kinetic barrier to reaction

High energy “Transition state”Intermediate between R & P

Products (P)

The energy barrier is critical for life

• Potentially deleterious reactions are blocked by EA

– Complex molecule degrading to simpler constituents

http://asm.wku.eduhttp://encyclopedia.quickseek.com/

DNAnucleotide

How do enzymes speed up reactions?

• New reaction pathway

• Lower activation energy

• Decreased energy barrier

2H2O2 → 2H2O + O2

Isolated: EA ~ 86 kJ/molIn the presence of catalase: EA ~ 1kJ/mol

Hydrogen peroxide

Binding of substrate to enzyme creates a new reaction pathway

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An enzyme changes EA NOT G

Affects RATE, not EQUILIBRIUM

Without enzyme

With enzyme

EA = G‡

How is EA lowered?

• Enzyme’s ‘goal’ is to reduce G‡

• Two ways enzymes can affect G

– Improve H– Improve S

EA =G‡ = H - TS

G‡ = Gtrans.state – Greactants

Enzymes alter the free energy of the

transition state

enthalpy entropy

-

Example: More favorable H

A B

AOHBH

A BH+

+ H2O

+OH-

+

Charge unfavorableUnstable transition st.

A BH+

Ionic interaction stabilizesthe positive charge

OH-

Example: More favorable S

Two moleculesMore ‘freedom’Higher disorder (high S)

One moleculeLower disorder (low S)Unfavorable entropically

ENZYME

Example: More favorable S

Enzyme/Reactant COMPLEX

Essentially a single molecule

ENZYME

Enzyme/Transition state complex

Still a single molecule

Not much difference entropically

Remember

1. Enzymes lower the energy barrier

2. Decrease EA (G‡)

3. Provide an environment where:

• Transition state is stabilized (lower enthalpy)• Change of disorder (entropy) is minimized

Enzymes create a new reaction pathway

Go vs. G‡ transition state vs. reaction intermediates rate limiting step

Factors contributing to enzyme catalysis

• Weak interactions between enzyme and transition state

• Transient covalent bonds between S and E

• Entropy optimization in ES complex formation

• Solvation shell surrounding S & E (entropy/hydrophobic interactions)

• Substrate distortion upon binding to noncomplementary E

• Proper alignment of catalytic functional groups

Common catalytic mechanisms

• General acid/base catalysis– Proton transfer– Reactions with charged

intermediates/AAs• Fumarase

– Precise positioning of acid/base: reaction occurs faster than specific acid/base reactions

• Free H+/OH-

Common catalytic mechanisms

• Covalent catalysis– Covalent bond formation

between E and S– Reaction path is altered

and new path has lower Ea

– Chymotrypsin (combination)

Common catalytic mechanisms

• Metal catalysis (metalloenzymes)– Ionic interactions

• Stabilize charged TS or orient charged substrate for reaction

• Carboxypeptidase

– Oxidation/reduction• Reversible changes in oxidation state of the metal• Electron transfer reactions • Transition metals• Catalase ezample