生物化學(xué)考研課件ch14酶動力學(xué)enzymekinetics

1、BiochemistryFor Speciality of Biotechnology or Bioengineering at CSUMr. XIA JinlanChapter 14 Enzyme kinetics Introduction to enzymes 酶（學(xué)）導(dǎo)論Factors affecting enzyme activity 影響酶活性的因素Enzyme assaysEnzyme kinetics and inhibition 酶動力學(xué)和抑制作用Introduction to enzymes 酶（學(xué)）導(dǎo)論The features of enzymes 酶的特征Some ter

2、ms of enzyme 酶的幾種術(shù)語Classes of enzymes 酶的種類Enzyme Nomenclature 酶的命名法Traditional nomenclature 傳統(tǒng)命名法Systematic nomenclature 系統(tǒng)命名法Expression of enzyme activity 酶活性的表示Components of enzyme 酶的成分Cofactors 輔助因子Powerful Catalytic power 強的催化力Increase reaction rate up to 1016 foldMake reaction taken place under

3、 relatively mild conditions of temperature and pHHigh specificity 高的特異性To the substrates that they act on and the products they formEnzyme activity can be regulated 酶活性可以被調(diào)控Basically are proteins, except few ribozymes 除了少量的核酶外,基本上是蛋白質(zhì) Enzymes have four basic features酶的四個基本特征Some terms of enzymes 酶的幾

4、種術(shù)語Proteoenzyme蛋白酶Ribozyme核酶Abzyme 抗體酶Holoenzyme全酶Apoenzyme脫輔酶Multimeric enzyme多聚合體酶(多亞基酶)Enzymes are classified into six classes based on the type of reaction catalyzed基于催化反應(yīng)的類型,酶可分為六種類型Oxidoreductases 氧化還原酶Transfer of electrons 電子的轉(zhuǎn)移Transferases 轉(zhuǎn)化(或轉(zhuǎn)移)酶Transfer of functional groups 分子間功能基團的轉(zhuǎn)移Hydr

5、olases 水解酶Hydrolysis reactions 水解反應(yīng)Lyases 裂解酶Cleavage of C-C, C-O, C-N etc., often forming a double bond C-C, C-O, C-N鍵的裂解,并通常形成雙鍵的反應(yīng)Isomerases 異構(gòu)酶Transfer of groups within a molecule 分子內(nèi)基團的轉(zhuǎn)移Ligases (or synthases) 鏈接酶(或合成酶)Bond formation coupled to ATP hydrolysis伴隨ATP水解的鍵合Enzymes are classified int

6、o six classes based on the type of reaction catalyzed NameType of reaction catalyzedExampleOxido-reductasesTransfer of electronsA- + B A + B-Alcohol dehydrogenaseTransfer-asesTransfer of functional groupsA-B +CA + B-CHexokinaseHydrolasesHydrolysis reactionsA-B + H2OA-H + B-OHTrypsinLyasesCleavage of

7、 C-C, C-O, C-N etc., often forming a double bondPyruvate decarboxylaseIsomerasesTransfer of groups within a moleculeMaleate isomaseLigases (or synthases)Bond formation coupled to ATP hydrolysisPyruvate carboxylaseA + B A-BX-A-B-Y Y-A-B-XX-A-B-Y A=B + Y-XTraditional Nomenclature of enzymesSystematic

8、nomenclature: Each enzyme is uniquely identified by a four-digit classification number Enzyme commission (EC) number: Four-digit classification numberWhat do these numbers signify? Trypsin EC 3.4.21.4Chymotrypsin EC 3.4.21.1Elastase EC 3.4.21.36Expression of enzyme activity 酶活性的表示 Initial rate (V0)

9、of the reaction mmol of substrate transformed per minute 每分鐘轉(zhuǎn)化底物的微摩爾數(shù)V0: X mmol.min-1 Enzyme unit 酶的單位, SI unitThe amount that catalyzes the formation of one mmol of product per minute 每分鐘催化1mmol產(chǎn)物的形成所需的酶量U: mmol.min-1Katal , SI unitAmount of enzyme catalyzing the conversion of one mole of substrate

10、 to product per second每秒鐘催化轉(zhuǎn)化1摩爾底物所需的酶量Katal: mol.s-1Expression of enzyme activity酶活性的表示 11Some expressions of the components of an enzyme酶的成分的幾種表示方式Enzyme 酶Protein part + Cofactors (Non-protein part) (蛋白部分+輔助因子(非蛋白部分) Holoenzyme全酶Apoenzyme + Prosthetic group (脫輔基酶+輔基)Cofactors 輔助因子Inorganic Inorgan

11、ic ion(s) 無機離子 Fe2+, Zn2+, Cu2+, etc.Inorganic cluster(s) 無機簇 Fe-S cluster, Mn-O complex,etcOrganic Coenzyme 輔酶 Which loosely bind to their enzymes Prosthetic group 輔基 Which tightly bind to their enzymesThe important coenzymes and prosthetic groups 重要的輔酶和輔基Coenzymes 輔酶Coenzyme A (CoA)NAD(H), NADP(H)

12、Nicotinamide adenine dinucleotide 煙酰胺腺嘌呤二核苷酸Nicotinamide adenine dinucleotide phosphateProsthetic groups 輔基FAD(H2), FMN(H2)*Flavin adenine dinucleotide 黃素腺嘌呤二核苷酸Flavin mononucleotide 黃素單核苷酸Biotin 生物素Heme 血紅素* FAD and FMN in most cases are prosthetic groups binding to flavin proteinsWater soluble vit

13、amins (Vitamin B)Thiamine (B1) 硫胺素In Coenzymes：thiamine pyrophosphate 存在于硫胺焦磷酸輔酶中Riboflavin (核黃素B2)As prosthetic group flavin protein: FAD and FMNNicotinic acid (niacin煙堿酸)In Coenzymes: NAD and NADPPantothenic acid (泛酸）In Coenzyme ABiotin (生物素）In prosthetic group: Biocytin生物胞素Lipid soluble vitamins

14、(Vitamin A, D, E, K)Vitamin ARetinal 視網(wǎng)膜素：vision, growth and reproductionMost coenzymes are derived from vitamins大多數(shù)輔酶源于維生素Some common coenzymes, their vitamin precursors and deficiency diseases Coenzyme/prosthetic groupPrecursorDeficiency diseaseCoenzyme APanthothenic acidDermatitis皮膚炎FAD, FMNRibof

15、lavin (Vitamin B2)Growth retardationNAD+ (Coenzyme I), NADP+ (Coenzyme II)Niacin (菸堿酸)Pellagra 玉蜀黍疹Thiamine pyrophosphateThiamine (Vitamin B1)Beriberi 腳氣病TetrahydrofolateFolic acidAnemia貧血癥Deoxyadenosyl cobalamin（脫氧腺苷鈷胺素）Cobalamin (Vitamin B12)Pernicious anemia惡性貧血癥Co-substrate in the hydroxylation

16、of proline in collagenVitamin C (ascorbic acid)Scurvy 壞血病Pyridoxal phosphate 磷酸吡哆醛Pyridoxine (vitamin B6)Dermatitis 皮膚炎Coenzyme ATransfer of acetyl groupNAD+, NADP+Coenzyme ofDehydrogenasesNAD(P)+ + H+ + 2 e-NAD(P)HReductionOxdationNAD+, NADP+H+ + 2 e-H-NAD(P)+ + H-NAD(P)HReductionOxdationGeneral re

17、actionFAD, FMNFig. 18.21, p591Isoalloxazine 異咯嗪Flavin mononucleotide, FMNFlavin adenine dinucleotide, FADRiboflavinAMPRibitol核醣醇FAD, FMNFAD + 2H+ + 2 e-FADH2ReductionOxdationFMN + 2H+ + 2 e-FMNH2ReductionOxdationChemical structure of biotinChapter 14 Enzyme kinetics Introduction to enzymes 酶（學(xué)）導(dǎo)論Fac

18、tors affecting enzyme activity 影響酶活性的因素Enzyme assaysEnzyme kinetics and inhibition 酶動力學(xué)和抑制作用Cofactors 輔助因子 (SEE the last section) pHTemperatureSubstrate concentration 底物濃度Product concentrationFactors affecting enzyme activity Chapter 14 EnzymesEffect of pHPepsin胃蛋白酶Papain木瓜蛋白酶Trypsin胰蛋白酶Cholinestera

19、se膽堿酯酶過氧化氫酶Effect of temperature50 C, decrease due to denaturation of enzymeBATypicallyEnzyme assays 酶的檢測Two most convenient waysDetg. the rate of appearance of product 測定產(chǎn)物形成的速率Detg. the rate of disappearance of substrate 測定反應(yīng)物減少的速率A linked enzyme assay way 酶聯(lián)檢測法First enzyme reaction + second enzym

20、e reactionMeasurements 測定Spectrophotometry 分光光度法 Enzyme electrode 酶電極法LDH: lactate dehydrogenase 乳酸脫氫酶COO-CHOHCH3+NAD+COO-COCH3Pyruvate丙酮酸Lactate乳酸+H+NADH+A340nmLDH葡萄糖氧化酶過氧化氫氧化酶葡萄糖酸Chapter 14 Enzyme kinetics Introduction to enzymes 酶（學(xué)）導(dǎo)論Factors affecting enzyme activity 影響酶活性的因素Enzyme assaysEnzyme

21、kinetics and inhibition 酶動力學(xué)和抑制作用Enzyme kinetics and inhibition (酶動力學(xué)和抑制作用)Catalytic mechanism 催化機制 Enzyme kinetics 酶動力學(xué)Michaelis-Menten modelEnzyme inhibition 酶的抑制Reversible inhibition 可逆抑制Nonreversible inhibition 非可逆抑制Enzymes catalyze chemical reaction through lowering its activation energyG0: not

22、 thermodynamically favorable Progress of reactionFree energy, GTransition state(uncatalyzed)Transition state(catalyzed)GuncatalyzedGcatalyzedSPGActivation Energy Rate Constant of Reaction Gk Arrhenius equationk =Ae-G/RTGkQuestions (self study) What does dynamic equilibrium mean?Enzymes can accelerat

23、e the attainment of the equilibrium position, can it change this position?Whats role of active site of an enzyme? By which forces the substrate binds to the active site of an enzyme?Chemical equilibrium: (self study)Michaelis-Menten modelHyperbolic curve雙曲線Double reciprocal or lineweaver-Burk plotV0

24、Time (min)Amount of product formed (mmol)Measurement and calculation of V0Enzyme inhibition 酶的抑制Inhibition 抑制的定義A phenomenon of lowering enzymes catalytic rate. Inhibitor 抑制劑Any molecule which acts directly on an enzyme to lower its catalytic rateTwo main types of inhibitionsIrreversible 非可逆抑制(抑制)Reversible 可逆(抑制)Competitive 競爭性Noncompetitive 非競爭性Mixed noncompetitive inhibition 混合型非競爭性Irreversible inhibition by covalent bin