Chapter 3. enzyme Composition Characteristics and mechanism

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1 Chapter 3. enzyme Composition Characteristics and mechanism
Kinetics of enzyme-catalzed reaction Regulation Names and classes Enzyme and medicine 2018/11/21

2 introduction biocatalyst生物催化剂: enzyme and ribozyme核酶
Enzyme---proteins having catalytic activity Ribozyme---nucleic acids having catalytic activity 2018/11/21

3 Enzyme is protein. 1926, Sumner美国,脲酶结晶，酶是蛋白质 J.B.Sumner
J.H.Northrop 1930年Northrop等得到了胃蛋白酶、胰蛋白酶和胰凝乳蛋白酶的结晶，进一步证明了酶是蛋白质 2018/11/21

4 ribozyme 1982年T.Cech等人发现四膜虫的rRNA前体能在完全没有蛋白质的情况下进行自我加工，发现RNA有催化活性
Thomas Cech University of Colorado at Boulder, USA 1982年T.Cech等人发现四膜虫的rRNA前体能在完全没有蛋白质的情况下进行自我加工，发现RNA有催化活性 1983年S.Altman等研究RNaseP（20%蛋白质和80%RNA），发现RNaseP中的RNA可催化E. coli tRNA的前体加工 Sidney Altman Yale University New Haven, CT, USA 2018/11/21

5 Section 1 molecular structure and function
Monomeric enzyme单体酶：one peptide chain Oligomeric enzyme寡聚酶：many subunits Multienzyme system多酶体系：complex of enzymes of different function. Multifunctional enzyme多功能酶 / Tandem enzyme串联酶: function different enzymes in one peptide chain. 2018/11/21

6 Ⅰ. Molecular composition of enzymes
inactive Simple enzyme单纯酶 enzyme apoenzyme酶蛋白 metal ion金属离子 Conjugated enzyme结合酶 cofactor辅助因子 Holoenzyme 全酶 active Small organic molecules　有机小分子 2018/11/21

7 Functions of metal ion:
Metalloenzyme金属酶:　bind metal ion tightly Metal activated enzyme金属激活酶: don’t bind metal ion tightly, but the metal ion is essential to the enzyme activity. Functions of metal ion: As active center, join in electron transfer,group tansfer and so on. Stablize steric structure of apoenzyme. Help the substrate binding. Neutralize anion, to lower static electricity repulsion of the reaction. 2018/11/21

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9 coenzyme辅酶 cofactor Prosthetic group辅基 2018/11/21

10 Ⅱ. Active center of enzymes
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11 Section 2. characteristics and mechanism of enzyme-catalyzed reaction
酶与一般催化剂的共同点common characters of enzymes and general catalysts： 反应前后没有质和量的变化the quality and mass of catalysts before the reaction are identical to that after reaction. 只能催化热力学允许的反应catalyze the reaction thermodynamics-permissive only. 只能加速可逆反应的进程，而不能改变反应的平衡点Accelerate reversible reaction proceeding, but can’t change the point of balance. 2018/11/21

12 Ⅰ. Characteristics of enzyme-catalyzed reaction
High efficiency 比非催化反应高108～1020倍； 可比一般催化剂高106～1013倍 尿素水解 脲酶／Ｈ＋：７１０１２ 苯酰胺水解 α-胰凝乳蛋白酶／Ｈ＋：6 106 25℃时，活化能每减少4.18kJ/mol,反应速度增加5.4倍． 2018/11/21

13 The enzyme-catalyzed reaction can be regulated可调节性
Specificity专一性 Absolute specificity: 一种底物 Relative specificity：一类底物 Stereospecificity：对立体异构体有选择性 The enzyme-catalyzed reaction can be regulated可调节性 2018/11/21

14 Ⅱ. Mechanism of enzyme-catalyzed reaction
Induced-fit hypothesis for forming of enzyme-substrate complex 　　酶在与底物相互接近时，其结构相互诱导、相互变形和相互适应，进而相互结合。 2018/11/21

15 Reaction mechanism 邻近效应proximity effect与定向排列orientation arrange
多元催化multielement catalysis与协同作用cooperation 表面效应surface effect 2018/11/21

16 Section 3. kinetics of enzyme-catalyzed reactions
Ⅰ. Influence of substrate concentration on reaction velocity 当底物浓度很小时，为一级反应．V= k [S] 随底物浓度逐渐增大，变为混合级反应 当底物浓度很大时，为零级反应.v= k. 2018/11/21

17 1. Michaelis equation米－曼氏方程式
1913, L. Michaelis and M.L.Menten 2018/11/21

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21 Km equal to [S] when the velocity is half of the Vmax. (mol/L)
2. Meaning of Km and Vm Km equal to [S] when the velocity is half of the Vmax. (mol/L) Km的大小反映该酶对底物亲和力的大小the value of Km weigh the affinity of enzyme with substrate. Km= K2+K3 K1 Km= K2 K1 = KS = [E] [S] [ES] If K2 >> K3, Km值是酶的特性常数，只与酶的结构、底物和反应环境有关，与酶的浓度无关。在10-6~10-2mmol/L之间。 2018/11/21

22 Vm, Vmax, 最大反应速度，酶完全被底物饱和时的反应速度．
K3, 转换数 turnover number: 　　当酶被底物充分饱和时，单位时间内每个酶分子催化底物转变为产物的分子数．大多数在1~104/s之间． K3 = Vm [E] 2018/11/21

23 3. Determination of Km and Vm
1 V = Km Vm [S]  + 1 V Km Vm r = 1 Vm 1 Km - 1 [S] 2018/11/21

24 Ⅱ. influence of enzyme concentration
v = k3 [E], as [S]>>[E] 2018/11/21

25 Ⅲ. Influence of temperature
Optimum temperature最适温度，most of them are in the range from 35 to 45℃ . 2018/11/21

26 Ⅳ. Influence of pH Optimum pH最适pH 常为中性，胃蛋白酶为1.8, 肝精氨酸酶为9.8． 2018/11/21

27 Ⅴ. Influence of inhibitors
the matters which decrease enzyme activity, but don’t make the enzyme denatured. Irreversible inhibition Reversible inhibition可逆性抑制 competitive inhibition竞争性抑制 non-competitive inhibition非竞争性抑制 uncompetitive inhibition反竞争性抑制 2018/11/21

28 1. Irreversible inhibition
the inhibitor combine with essential group of enzyme active center by covalent bond, resulting in enzymatic activity loss. 2018/11/21

29 2. Reversible inhibition
the inhibitor combine with enzyme,or enzyme-substrate complex by non-covalent bond, resulting in enzymatic activity decrease or loss. the inhibitor can be removed by dialysis, or ultrafiltration competitive inhibition竞争性抑制 non-competitive inhibition非竞争性抑制 uncompetitive inhibition反竞争性抑制 2018/11/21

30 (1) competitive inhibition
the structure of inhibitor is similar to substrate. This make it combine with enzyme competively to substrate. 2018/11/21

31 Characteristics of competitive inhibition
Inhibitor structure is similar to that of substrate Degree of inhibition is determined by their affinity and substrate concentration. Vmax is invariable, apparent Km increase. 2018/11/21

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34 (2)non-competitive inhibition非竞争性抑制
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35 Characteristics of non-competitive inhibition
Inhibitor combine with essential groups out of active center, no competitive . Degree of inhibition is determined by the concentration of inhibitor. Vmax decrease, Km is invariable 2018/11/21

36 (3)uncompetitive inhibition反竞争性抑制
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37 Characteristics of uncompetitive inhibition
Inhibitor combine with enzyme-substrate complex only. Degree of inhibition is determined by concentration of both inhibitor and substrate. Vm decrease, Km decrease. 2018/11/21

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39 Ⅵ. Influence of activator
Activator激活剂：substance which change enzymes inactive into active state, or which increase enzyme activity, such as Mg2+, K+, Cl-. Essential activator Non-essential activator 2018/11/21

40 Ⅶ. Determination of enzyme activity ; enzyme activity unit
The enzyme activity is determined by the reaction ratio. 1国际单位 IU = 在特定条件下，每分钟催化1μmol底物转化为产物所需要的酶量．Quantity of enzyme which can catalyze 1μmol substrate change into product at certain condition. 1催量kat＝在特定条件下，每秒钟催化1mol底物转化为产物所需要的酶量． 1IU =  10-9 kat 2018/11/21

41 Section 4. the regulation of enzyme
Regulated target: key enzyme Ways of regulation: Regulation of enzyme activity (fast). Regulation of enzyme quantity (slow). 2018/11/21

42 Ⅰ. Regulation of enzyme activity
1.Zymogen and activation of zymogen酶原与酶原激活 Zymogen: 有些酶在细胞内合成或刚分泌时，无催化活性，这种无催化活性的酶的前体． activation of zymogen： the process of inactive zymogen change into active enzyme, also the process of forming or exposing active center. 2018/11/21

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44 2. Allosteric regulation变构调节
效应剂 2. Allosteric regulation变构调节 调节亚基 催化亚基 变构酶 2018/11/21

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46 3. Regulated by covalent modification of enzyme
磷酸化phosphorylation  脱磷酸化dephosphorylation 乙酰化acetylation  脱乙酰化产deacetylation 甲基化methylation  脱甲基化demethylation 腺苷化adenosination  脱腺苷化deadenosination RSH  RS-SR 2018/11/21

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48 Ⅱ. Regulation of enzyme quantity
Induce or repress synthesis of apoenzyme Regulation of enzyme degradation 2018/11/21

49 Ⅲ. Isoenzyme同工酶 Isoenzyme: enzymes which catalyze same reaction, but their molecule structure, physico-chemical property　and immunology charcter are different.they must be the enzymes encoded by different genes or mRNA, exist in different tissues of a body,or in different subunits of a cell. 2018/11/21

50 Section 5. naming and classification of enzyme
Ⅰ. 命名 习惯名称推荐名称 系统命名法： 底物：底物　反应性质． 2018/11/21

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52 Ⅱ. classification 氧化还原酶oxidoreductases 转移酶transferases 水解酶hydrolases
裂解酶lyases：催化从底物移去一个基团并留下双键的的反应或其逆反应． 异构酶isomerases 合成酶ligases 2018/11/21

53 Section 6. relation of enzyme with medicine
Ⅰ. Enzyme and disease Enzyme and diseases occurrence Enzyme and diseases diagnosis Enzyme and diseases treatment 2018/11/21

54 Ⅱ. Application of enzyme in medicine
used in clinical test as reagent Used in clinical treatment as medicine Used in scientific research and production 工具酶 　酶标记测定法 　固定化酶 　抗体酶 2018/11/21

55 选择题练习 酶化学 2018/11/21

56 E．酶是由活细胞合成具有催化作用的蛋白质
1. 关于酶概念的叙述下列哪项是正确的？ A．所有蛋白质都有酶的活性 B．其底物都是有机化合物 C．其催化活性都需要特异的辅助因子 D．体内所有具有催化活性的物质都是酶 E．酶是由活细胞合成具有催化作用的蛋白质 2018/11/21

57 2．关于酶活性中心的叙述下列哪项是正确的？
A．所有酶的活性中心都有金属离子 B．所有的抑制剂都作用于酶的活性中心 C．所有的必需集团都位于酶的活性中心 D．所有酶的活性中心都含有辅酶 E．所有的酶都有活性中心 2018/11/21

58 3. 酶加速化学反应的根本原因是( ) A. 升高反应温度 B. 增加反应物碰撞频率 C. 降低催化反应的活化能 D. 增加底物浓度
3. 酶加速化学反应的根本原因是( ) A. 升高反应温度 B. 增加反应物碰撞频率 C. 降低催化反应的活化能 D. 增加底物浓度 E. 降低产物的自由能 2018/11/21

59 A. Complex of enzyme with substrate
4. Holoenzyme refer to ( ) A. Complex of enzyme with substrate B. Complex of enzyme with suppressant C. Complex of enzyme with cofactor D. Inactive precursor of enzyme Complex of enzyme with allosteric effector 2018/11/21

60 5. 金属离子作为辅助因子的作用错误的是( ) A. 作为酶活性中心的催化基团参加反应 B. 与稳定酶的分子构象无关
5. 金属离子作为辅助因子的作用错误的是( ) A. 作为酶活性中心的催化基团参加反应 B. 与稳定酶的分子构象无关 C. 可提高酶的催化活性 D. 降低反应中的静电排斥 E. 可与酶、底物形成复合物 2018/11/21

61 6. 活化能的概念是指( ) A. 底物和产物之间能量的差值 B. 参与反应的分子所需的总能量 C. 分子由一般状态变成活化态所需能量
6. 活化能的概念是指( ) A. 底物和产物之间能量的差值 B. 参与反应的分子所需的总能量 C. 分子由一般状态变成活化态所需能量 D. 温度升高时产生的能量 E. 以上都不是 2018/11/21

62 7. 酶促反应动力学所研究的是( ) A. 酶的基因来源 B. 酶的电泳行为 C. 酶的诱导契合 D. 酶分子的空间结构
7. 酶促反应动力学所研究的是( ) A. 酶的基因来源 B. 酶的电泳行为 C. 酶的诱导契合 D. 酶分子的空间结构 E. 影响酶促反应速度的因素 2018/11/21

63 8. Michaelis-Menten enzyme kinetics diagram of curves is a ( )
A. straight line B. rectangular hyperbola C. S shape curve D. parabola E. Not above all 2018/11/21

64 9. 关于Km的意义正确的是( ) A. Km为酶的比活性 B. 1/Km越小,酶与底物亲和力越大 C. Km的单位是mmol/min
D. Km值是酶的特征性常数之一 E. Km值与酶的浓度有关 2018/11/21

65 10. 竞争性抑制剂的特点是( ) A. 抑制剂以共价键与酶结合 B. 抑制剂的结构与底物不相似 C. 当抑制剂的浓度增加时,酶变性失活
10. 竞争性抑制剂的特点是( ) A. 抑制剂以共价键与酶结合 B. 抑制剂的结构与底物不相似 C. 当抑制剂的浓度增加时,酶变性失活 D. 当底物浓度增加时,抑制作用不减弱 E. 抑制剂和酶活性中心外的部位结合 2018/11/21

66 11. In anticompetitive inhibition of enzyme, the reaction kinetics parameter change as ( )
Km↑,Vmax invariably Km↓,Vmax↓ Km invariably,Vmax↓ D. Km↓,Vmax invariably E. Km↓,Vmax↑ 2018/11/21

67 12. 有机磷农药与酶活性中心结合的基团是( ) A. 组氨酸上的咪唑基 B. 赖氨酸上的ε-氨基 C. 丝氨酸上的羟基
12. 有机磷农药与酶活性中心结合的基团是( ) A. 组氨酸上的咪唑基 B. 赖氨酸上的ε-氨基 C. 丝氨酸上的羟基 D. 半胱氨酸上的巯基 E. 谷氨酸上的γ-羧基 2018/11/21

68 B. 如底物与一亚基结合后,使其他亚基迅速与底物结合程正协同效应
13. 关于变构酶的论述错误的是( ) A. 变构酶为多亚基组成 B. 如底物与一亚基结合后,使其他亚基迅速与底物结合程正协同效应 C. 正协同效应的底物浓度曲线呈矩形双曲线 D. 底物与一亚基结合后,使其亚基结合底物能力减少称负协同效应 E. 变构效应剂与一亚基结合后,使酶其他亚基迅速与底物结合为异促协同效应 2018/11/21

69 14. –SH is one enzyme’s essential group
–SH is one enzyme’s essential group. Which substance can protect this enzyme from oxidation? A. Cys B. GSH C. urea D. ionic detergent E. ethanol 2018/11/21

70 15. 快速调节可通过( ) A. 磷酸化与去磷酸化 B. 腺苷酸化与腺苷酸化 C. 变构调节 D. 改变酶的合成速度
15. 快速调节可通过( ) A. 磷酸化与去磷酸化 B. 腺苷酸化与腺苷酸化 C. 变构调节 D. 改变酶的合成速度 E. 酶促反应的可调节性 2018/11/21

71 16. The characteristic constants of enzymes include ( )
A. Enzymic optimum temperature B. Enzymic optimum pH C. Vmax D. Km E. KS 2018/11/21

72 D. 抑制程度强弱取决于药物与酶底物浓度的相对比例
17. 磺胺药的抑菌机理是( ) A. 竞争性抑制二氢叶酸合成酶的活性 B. 干扰体内核酸的代谢 C. 结构与二氢叶酸相似 D. 抑制程度强弱取决于药物与酶底物浓度的相对比例 E. 磺胺药是二氢叶酸合成酶的变构抑制剂 2018/11/21

73 18. Cofactors of enzyme are ( )
A. Micromolecule organic compounds B. metal ion C. vitamine D. various kinds of organic and inorganic compounds E. A kind of conjugated protein 2018/11/21

74 19. 某种酶的活性依赖于酶活性中心的必需基团-SH,能保护此酶不被氧化的物质是( )
A. GSH B. 维生素C C. 半胱氨酸 D. 维生素A E. 两价阳离子 2018/11/21

75 20. 酶分子上必需基团的作用是( ) A. 与底物结合 B. 催化底物发生化学反应 C. 含砷的有机化合物 D. 决定辅酶结构
20. 酶分子上必需基团的作用是( ) A. 与底物结合 B. 催化底物发生化学反应 C. 含砷的有机化合物 D. 决定辅酶结构 E. 维持酶分子空间结构 2018/11/21

76 Thank you! 2018/11/21