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Enzymes II Dr. Kevin Ahern Enzymes II Dr. Kevin Ahern

Enzymes II Dr. Kevin Ahern - PowerPoint Presentation

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Enzymes II Dr. Kevin Ahern - PPT Presentation

E S ltgt ES ltgt ES ltgt EP ltgt E P MichaelisMenten Kinetics E S ltgt ES ltgt ES ltgt EP ltgt E P Rate of Formation Substrate Product Enzyme Enzyme ID: 688926

enzymes enzyme michaelis substrate enzyme enzymes substrate michaelis menten kinetics max high product time considerations kinetic change cat state

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Presentation Transcript

Slide1

Enzymes II

Dr. Kevin AhernSlide2

E+S <=> ES <=> ES* <=> EP <=> E+PSlide3

Michaelis-Menten Kinetics

E+S <=> ES <=> ES* <=> EP <=> E+P

Rate of FormationSlide4

Substrate

Product

Enzyme

Enzyme-

Substrate

Complex

Michaelis-Menten KineticsSlide5

Michaelis-Menten Kinetics

Substrate high

Enzyme high

ES low

Product lowSlide6

Kinetic Considerations

Pre-steady state

[E] and [ES]

varying widely

Steady state

[E] and [ES]

relatively constantSlide7

Pre-steady State Kinetics

Can give info on reaction mechanism, rate of ES formationSlide8

Enzymes

Kinetic ConsiderationsSlide9

Enzymes

Kinetic Considerations

Initial Velocity (V

0

) - Measured as [Product]/Time

Low [S], Low V

0

(Enzymes Often Idle)

Substrate Concentration (Molarity)

High [S], High V

0

(Enzymes Almost Always Busy)Slide10

Kinetic ConsiderationsSlide11

Michaelis-Menten Kinetics

Parameters

K

m

K

m

= Substrate Concentration that Gives V

max

/2

NOT Vmax/2Slide12

Michaelis-Menten Kinetics

Considerations

v

0

=

V

max

[S]

K

m

+ [S]

Enzymes That Don’t Follow Michaelis-Menten Kinetics Include Those That Bind Substrate Cooperatively - Binding of One Substrate Affects Binding of Others Slide13

Control of Enzyme Activity

Allosteric Control

Substrate Does Not Change Enzyme

Binding of Substrate

Substrate Does Change Enzyme

Binding of SubstrateSlide14

Michaelis-Menten Equation

V

max

occurs when an enzyme is saturated by substrate

V

max

varies with amount of enzyme usedKm is a measure of an enzyme’s affinity for its substrateKm value inversely related to affinity

High K

m

= Low Affinity

Low K

m

= High AffinitySlide15

Michaelis-Menten Kinetics

V

max

and K

cat

V

max

[Enzyme Used]

=

[Product]

[Enzyme Used] *Time

V

max

is Proportional to the Amount of Enzyme Used

in an Experiment - Not Useful for Comparing Enzymes

The Two Concentrations Cancel Out.

The Result is a Number Per time (say 1000/second).

It is the Number of Product Molecules Made by Each Enzyme Molecule Per Time.

It is Also Known as the Turnover Number or K

cat

and

Does Not Vary with the Amount of Enzyme

Since V

max

is a Velocity, and Velocity = [Product]/Time,Slide16

Perfect Enzymes

Maximum K

cat

/K

MMutation leads to reduced Kcat/KMDiffusion of substrate limitingSlide17

Triose Phosphate IsomeraseSlide18

Enzyme Co-factorsSlide19

Michaelis-Menten Kinetics

Lineweaver Burk Plot

Also Called Double Reciprocal Plot

Uses Same Data as V vs. [S] Plot,

but Inverts All Data for the Plotting

Linear for Enzymes Following Michaelis Menten Kinetics

Direct Reading of -1/K

m

and 1/V

maxSlide20

RibozymesSlide21

Metabolic Melody

You know it's true

So when an effector fits

It will just rearrange

all the sub-u-nits

Inside an

ENZYME!

Flipping from R to T

ENZYME!

Slow catalytically

ENZYME!

No change in Delta G

You should relax

When seeking out the Vmax though

There are many steps

Enzymes

Lineweaver Burk Can save a scientist work

With just two intercepts Enzymes  

Plotting all the data from kinetic exploration

Lets you match a line into a best fitting equation Here's what you doBoth axes are inverted then

You can determine Vmax and Establish Kmfor your ENZYMES!Sterically holding tight

ENZYMES!

Substrates positioned right

ENZYMES!

Inside the active site

Enzymes (Enzymes, enzymes, enzymes)

Enzymes

(To the tune of "

Downtown

")

Copyright

Kevin Ahern

Reactions alone

Could starve your cells to the bone

Thank God we all produce

Enzymes

Units arrange

To make the chemicals change

Because you always use

Enzymes  

Sometimes mechanisms run like they are at the races

Witness the Kcat of the carbonic anhydrases

How do they work?

Inside of the active site

It just grabs onto a substrate

and squeezes it tight

In an

ENZYME!

CAT-al-y-sis

In an

ENZYME!

V versus S

In an

ENZYME!

All of this working for you

Energy peaks

Are what an enzyme defeats

In its catalysis

Enzymes

Transition state

Is what an enzyme does great

And you should all know this

Enzymes

Catalytic action won't run wild - don't get hysteric

Cells can throttle pathways with an enzyme allosteric