Enzymes

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Enzymes

• Biological catalysts

• Made primarily from proteins

• Bind to substrate

• Not used up

• Speed up reaction

CatalysisUncatalyzed

G

time

R

P

Transition

EA

time

R

P

G

Catalyzed

Catalysts

• Increase rate of reaction

• Provide activation energy– Energy is conserved– Activation energy has to come from

somewhere

Sources of Activation energy

• Binding energy– Solvent released from active site when

substrate binds increases ΔSsurr

• Induced fit– Enzyme forces substrate into unstable

transition state– Catalytic antibodies

• Binding substrate brings reactive groups together

Catalysis by an enzyme

• E + S > ES > EP > E + P

Enzyme Example Serine Proteases

• http://www.bio.cmu.edu/courses/03231/Protease/SerPro.htm

• Active site and substrate– Note side chain

interactions– Substrate diffuses into

active site

Serine proteases

• Substrate binds to active site

• Chemical groups interact

• Force substrate into unstable intermediate

Serine Proteases

• Peptide bond is cleaved

• Serine in active site is bound to carboxyl side of peptide

Serine proteases

• Half of protein diffuses out

• Enzyme used up– Has to be regenerated

Serine proteases

• Water diffuses into active site

• Juxtaposes chemical groups – Similar reaction to first

Serine proteases

• New unstable intermediate generated

Serine Proteases

• Carboxyl end of peptide diffuses out

• Enzyme regenerated • Ready for another

round

Serine protease summary

• Enzyme stabilized by various side chain interactions

• Substrate binds to enzyme.– Fits pocket– Forms unstable intermediate– Chemical groups on enzyme do reaction

• Chemistry happens and product diffuses out

• Enzyme regenerated

Kinetics

• Study of reaction rates• Why?

– Used to determine mechanisms

• Michaelis Menton kineticsV = rate of reaction

• Vmax = maximum reaction rate

[S]• Substrate concentration• Km = substrate concentration where rate is half maximal

Michaelis-Menton Plot

Enzyme inhibition, Competitive

• Both substrate and inhibitor bind to active site– Compete

• Inhibitor blocks substrate from binding

E S

I

EI

S

Michaelis- Menton Plot for competitive inhibition

uninhibVmax

Vmax/2

Km

inhibited

Kmi

V

[S]

Michaelis- Menton Plot for competitive inhibition

• Vmax is not changed

• Km increased

– Since substrate has to outcompete the inhibitor for the active site, it takes more substrate to get to the same rate.

Noncompetitive Inhibition

S

I

• Inhibitor binds to an allosteric site on the enzyme• Changes active site so substrate doesn’t bind

I

Michaelis- Menton Plot for noncompetitive inhibition

uninhib

Vmax/2

Km

Vmax uninhib

Vmax inhib

V

[S]

Michaelis- Menton Plot for noncompetitive inhibition

• Inhibition lowers Vmax

• Km unchanged

– Since inhibitor doesn’t bind to active site, changing amount of substrate will have no effect

Michaelis Menton Kinetics Summary

• Competitive inhibition– Inhibitor competes with substrate for active

site• Vmax unchanged• Km increased

• Noncompetitive inhibition– Inhibitor binds to allosteric site changing

active site• Vmax lowered• Km unchanged

Allosteric Activation

• Active site will not bind substrate• Allosteric activator binds and changes shape of

active site• Now substrate binds

S

A

A

S