Transcript
Page 1: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics

酶 Enzymes

Page 2: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics

contents

• Introduction of enzymes

• Catalytic Mechanisms of enzymes

• Enzymatic reactionkinetics

Page 3: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics

Enzymes are biocatalysts with high

specificity and highly catalytic efficency

produced by living cells.

Besides proteins, ribozymes and

deoxyribozymes are also enzymes.

What is enzyme?

Ⅰ Introduction of enzymes

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( 1 ) producing site : living cells

( 2 ) property : most——proteins

a few——RNA DNA

( 3 ) function : biochemical catalyzation

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Structure and Function of Enzymes

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Structure of enzymes

enzymes

Simple enzymes——proteins

apoenzyme

prosthetic group

coenzyme

Conjugated enzymes(holoenzyme)

cofactor

1.Composition of Enzyme Molecules

Holoenzyme = Apoenzyme + Cofactors

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2. Active site of an enzyme

It is an area of enzyme;It is consist of a few amino acids residues;The residues are closed to each other in space;It combines substrate and catalyzes a reaction.

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substrate Essential groups

outside of active site

Binding groups

catalytic group

active site

Active site of enzymes

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Primary structure and spatial structure of chymotrypsin

3. Structure of enzyme

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Three-dimensional structure Substrate binding

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Ⅱ Properties and Catalytic Mechanisms of Enzymes

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2. Highly Specificity of Enzymes

3. Activities of enzymes can be regulated

1. Highly Catalytic Activity of Enzymes

Properties of Enzymes

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Catalytic Mechanisms of enzymes

Enzymes accelerate reactions by decreasing the activation energy.

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For a biochemical reaction to proceed, the energy barrier needed to transform the substrate molecules into the transition state has to overcome. The transition state has the highest free energy in the energy pathway.

Formation of Enzyme-substrate Complex and Induced-fit Hypothesis

The combination of substrate and enzyme forces the substrate to become the transition state.

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Energy diagram for catalyzed and uncatalyzed reactions .

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Enzyme-substrate complex

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Induced fit model

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Induced fit model

Active sites in

the uninduced

enzyme

Binding of the first substrate (gold)

induces a conformational shift that

facilitates binding of the second

substrate (blue), with far lower

energy than otherwise required.

When catalysis

is complete, the

product is

released, and

the enzyme

returns to its

uninduced

state.

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Hexokinase-D-glucose complexHexokinaseInduced fit

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(1) Proximity and orientation effects

Catalytic mechanisms of enzymes

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FIGURE 3-7 Proximity effects and orientation arrange

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(2) Electrostatic effects

The active sites of enzymes are often hydrophobic.

Water is largedly excluded from the active site because of

the low dielectric constant. This increases the

electrostatic interactions between enzymes and

substrates and accelerate velocity of the reaction.

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(3) Acid-base catalysis

Side chain groups in enzyme active sites, act as

proton donors or acceptors are termed general acids or

general bases.

The transition state is stabilized and the rate of

reaction is increased by adding or removing proton from

reactants. These chemical groups are imidazole group,

amino group, carboxyl group, and so on.

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(4) Covalent catalysis

The attack of nucleophilic

or electrophilic group in the

enzyme active site upon the

substrate results in covalent

binding of substrate to the

enzyme as an intermediate in

the cause of catalysis.

Ser-OH

—CH2—S· ·

:H

—CH2—O· ·

:H

Cys-SH

—CH2—C=CH

HN N

CH

:His-imidazole

nucleophilic

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Ⅲ Enzymatic reaction kinetics

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ⅠⅠEffect of Effect of concentration of substratesconcentration of substrates on on enzyme reaction speedenzyme reaction speed

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1.

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2. sense and application of Km( 米氏常数 )

sense sense ::

(( 11 )) KmKm ,, that is the concentration of substrate that is the concentration of substrate

when reaction speed reach to ½ V when reaction speed reach to ½ V maxmax. .

(( 22 )) Km can present the affinity between enzyme and Km can present the affinity between enzyme and

substrate.substrate.

(( 33 )) Km is the mark constant.Km is the mark constant.

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Application:Application:

(( 11 )) To be used to identify enzyme; To be used to identify enzyme;

(( 22 )) To estimate the best substrate of enzyme; To estimate the best substrate of enzyme;

(( 33 )) To calculate the concentration of substrate at the To calculate the concentration of substrate at the

fixed speed;fixed speed;

(( 44 )) To infer whether the enzyme is regulated by To infer whether the enzyme is regulated by

concentration of substrate;concentration of substrate;

(( 55 )) To estimate the trend of reaction.To estimate the trend of reaction.

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3. Calculation metheds of Km3. Calculation metheds of Km

1.1. 双倒数作图双倒数作图

林贝氏方程林贝氏方程

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V[S] Km

Vmax[S]

Vmax+=

1

-Km

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ⅡEffect of pH on enzyme reaction speed

1. affect the combination of enzyme 1. affect the combination of enzyme and substrateand substrate

2. affect the conformation of enzyme 2. affect the conformation of enzyme moleculemolecule

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动物体内多数酶的最适动物体内多数酶的最适 pHpH 接近中性 接近中性

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Ⅲ Ⅲ Effect of Effect of temperature temperature on enzyme reaction speedon enzyme reaction speed

10 20 30 40 50 60 70 80 900

20

40

60

80

100

Temperature OC

Rel

ativ

e A

ctiv

ity

(%)

最适温度

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Ⅳ Effect of concentration of enzyme on reation speed

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activator:activator: the substance that could the substance that could

increase the activity of enzyme and increase the activity of enzyme and

elevate the enzyme reaction speed.elevate the enzyme reaction speed.

Ⅴ Effect of activator on enzyme reation speed

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Inhibitor: Inhibitor:

the substance could decrease the the substance could decrease the

activity of enzyme.activity of enzyme.

Ⅵ Effect of inhibitor on enzyme reation speed

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Types of inhibitingTypes of inhibiting

1. irreversible inhibition1. irreversible inhibition

specifity inhibitionspecifity inhibition

non-specifity inhibitionnon-specifity inhibition

2. reversible inhibition2. reversible inhibition

competitive inhibitioncompetitive inhibition

noncompetitive inhibitionnoncompetitive inhibition

uncompetitive inhibitionuncompetitive inhibition

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non-specifity inhibitionnon-specifity inhibition

Cl

As

Cl

CH CHCl + E

SH

SHE

S

As

S

CH CHCl + 2HCl

1. irreversible inhibition1. irreversible inhibition

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specifity inhibitionspecifity inhibition

R O

R' O

P

O

X

+ HO E

R O

R' O

P

O

O E

+ HX

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the inhibitor was attached to enzyme with the inhibitor was attached to enzyme with

non covalent bond to decrease or lose the non covalent bond to decrease or lose the

activity of enzyme, but the inhibitor could be activity of enzyme, but the inhibitor could be

remove by dialysis.remove by dialysis.

2. reversible inhibition2. reversible inhibition

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competitive inhibitioncompetitive inhibition

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Page 46: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics

双倒数作图双倒数作图

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Characteristic of competitive inhibitionCharacteristic of competitive inhibition

⑴ ⑴ 抑制剂往往是酶的抑制剂往往是酶的底物结构类似物底物结构类似物

⑵ ⑵ 抑制剂、底物与酶的抑制剂、底物与酶的结合部位相同结合部位相同—— —— 酶 酶

的活性中心的活性中心

⑶ ⑶ 抑制作用可以被抑制作用可以被高浓度的底物高浓度的底物减低以致消除减低以致消除

⑷ ⑷ (( 表观)表观) KmKm 值增大,值增大, VmVm 值不变值不变

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noncompetitive inhibitionnoncompetitive inhibition

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双倒数作图双倒数作图

Page 54: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics

⑴ ⑴ 非竞争性非竞争性抑制剂结构不一定与底物抑制剂结构不一定与底物的的结构类似结构类似;;⑵ ⑵ 抑制剂与酶的抑制剂与酶的活性中心外的位点活性中心外的位点结合;结合;⑶ ⑶ 底物浓度的改变对抑制程度无影响,底物浓度的改变对抑制程度无影响,抑制程度取抑制程度取决于抑制剂的浓度;决于抑制剂的浓度;

⑷ ⑷ 动力学参数:动力学参数: KmKm 值不变,值不变, (( 表观表观 )Vm)Vm 值降低值降低。 。

Characteristic of noncompetitive inhibitionCharacteristic of noncompetitive inhibition

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uncompetitive inhibitionuncompetitive inhibition

Page 56: 酶 Enzymes. contents Introduction of enzymes Catalytic Mechanisms of enzymes Enzymatic reactionkinetics
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双倒数作图双倒数作图

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(( 11 ) 反竞争性抑制剂只与酶) 反竞争性抑制剂只与酶————底物的中间产底物的中间产

物结合,不与单独的酶结合。物结合,不与单独的酶结合。

(( 22 )动力学参数:)动力学参数: (( 表观表观 )Km)Km 、、 VmaxVmax 都变都变

小。小。

(( 33 )抑制程度与)抑制程度与 [S] [S] 和和 [ I ][ I ] 成正比。成正比。

Characteristic of uncompetitive inhibitionCharacteristic of uncompetitive inhibition

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comparison of comparison of reversible inhibitionreversible inhibition


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