Chapter 3 Enzyme1 introduction to enzymes

1. Chapter 3 Enzyme1introduction to enzymes

2. Lecture Outline • Introduction to enzyme • Properties and catalytic mechanisms of enzymes • Structure and function of enzymes • Nomenclature and classification of enzyme • Kinetics of enzyme-catalyzed reaction • Regulation of enzyme activity • Clinical application of enzymes

3. Essential Question: What are enzymes, and what do they do? • Living Cells: Thousands of chemical reactions are proceeding very rapidly at any given instant. • All reactions are catalyzed by special biocatalysts, enzymes —protein and occasionally RNA. • Reactions in cells are different from general chemical reactions: easily, rapidly regulated and controlled • Enzyme-catalyzed reaction take place usually under relatively mild conditions.

4. Snail without enzyme catalyst Snail with enzyme catalyst What do enzymes do? • Enzymes are biological catalysts that accelerate the rates of chemical reactions.

5. What is the difference between an enzyme and a protein? Protein RNA Enzymes All enzymes are proteins except some RNAs and not all proteins areenzymes

6. Enzyme Product Substrates Substrates, products, and enzymes Enzymes catalyze the rate at which substrates are converted to product

7. Enzymes catalyze the conversion of substrates into products • What is a substrate? • A substrate is the compound that is converted into the product in an enzyme catalyzed reaction. • For the reaction catalyzed by aldolase, fructose 1,6-phosphate is the substrate.

8. What is the difference between enzyme catalyzed reactions and uncatalyzed chemical reactions? • Enzyme catalyzed reactions are much faster than uncatalyzed reactions. • Enzyme catalyzed reactions display saturation kinetics with respect to substrate concentration. • Enzyme catalyzed reactions are optimized for specific temperature and pH values.

9. Molecular components • Simple enzymes:enzymes require no other chemical groups other than their amino acid residues for activity. consists of only one peptide chain. • trypsin, chymotrypsin, ribonuclease A • Conjugated enzymes:enzymes contain chemical groups other than AA, the non-amino acid parts are usually called cofactors • the protein part alone called apoprotein(apoenzyme). • holoenzyme= apoenzyme (pr) +cofactor (non-pr)

10. Many vitamins, organic nutrients required in small amounts in the diet, are precursors of cofactors. • Cofactors: metal ions, small molecules, • Cofactors are divided two groups according to the binding ability. • ----coenzyme • -----prosthetic group

11. Coenzymes: • Loosely bind to apoenzyme. Be able to be separated with dialysis • Accepting H+ or group and leaving to transfer it to others, or vise versa. • Prosthetic groups: • Tightly bind through either covalent or many non-covalent interactions. • Remained bound to the apoenzyme during course of reaction.

12. Metal ions • Metal-activatedenzyme: ions necessary but loosely bound. Often found in metal-activated enzyme. Metal ion is essential for activity • Metalloenzymes: ions tightly bound • Particularly in the active center, transfer electrons, bridge the enzyme and substrates, stabilize enzyme conformation, neutralize the anions. • Metal ion is retained throughout purification

13. Organic componds • Small size and chemically stable compounds • Transferring electrons, protons and other groups • Vitamin-like or vitamin-containing molecule • Coenzymes often function as transient carriers of specific (functional) groups during catalysis.

14. Monomeric\oligomeric\multienzyme enzymes • Monomeric enzyme: only contain a polypeptide chain with tertiary structure. • Oligomeric enzyme: contain two or more polypeptide chains associated by noncovalent forces. • Multienzyme complex: different enzymes catalyzed sequential reactions in the same pathway are bound together. • Multifunctional enzyme(tandem enzyme): a single polypeptide chain with multiple activities

15. Components of enzyme • Essential groups: all groups essential for maintaining the enzyme activity • active center: • a specific region of the enzyme that contains some chemical groups for binding substrates and transforming it into products.( some functional groups are close enough in space to form a portion) • The active site is a three-dimensionalconformation formed by groups that come from different parts of the linear amino acid sequence. Look like a cleft or a crevice. • hydrophobic.

16. Two essential groups • Active site contains • binding groups (specificity of binding): to associate with the reactants to form an enzyme-substrate complex • determine specificity for substrate • catalytic groups (participate in the catalytic processes) :to catalyze the reactions and convert substrates into products • determine characteristics of catalyzation

17. Maintain conformation of enzyme Substrate Other essential groups Ser: OH Cys: SH His: imidazole Catalytic groups Binding groups Active site

18. Essential groups of the active site

19. In many enzymes, the active site has shape complementary to those of their substrates only after the substrates are bound (the induced fit). • For conjugated enzymes, the active site always contains coenzymes.

20. Two models have been proposed for substrate and enzyme binding: (1)Lock and key model (2) Induced fit model

21. Emil Ficher: (1894) • The ‘Lock & Key’ hypothesis • - explains substrate specificity • - says nothing about why catalysis occurs • Dan Koshland: (1958) • ‘Induced Fit’ hypothesis: • - enzymes prefer to bind to a distortion of the substrate that resembles the transition state • - both enzyme and substrate must adjust to one another • - in reality, enzyme is not ‘distorted’ but has evolved to bind in a certain way and sometimes undergo conformational changes

22. Lock and key model

23. Induced fit model

24. An Example: Induced conformational change in hexokinase Catalyzes phosphorylation of glucose to glucose 6-phosphate during glycolysis such a large change in a protein’s conformation is not unusual BUT: not all enzymes undergo such large changes in conformation

25. Advantage of the induced fit mechanism • The active site can be open to allow substrates to bind, then close over the substrates to provide optimum transition state stabilization Disadvantage of the induced fit mechanism • Energy that would otherwise be used to help stabilize the transition state of the reaction • Energy must be used to induce the conformational change in the enzyme.

26. Notice that in both the Lock and Key and Induced Fit models, the active site is designed to stabilize the transition state of the reaction.

27. Several factors contribute to enzyme catalysis • Proximity and orientation effects • Electrostatic effects • Acid-base catalysis • Covalent catalysis

28. What characteristic features define enzymes? • Enzymes are remarkably versatile biochemical catalysts that have in common three distinctive features: catalytic power, specificity, and regulation • Common features: 2 “do” and 2 “don’t • Unique features: 3 “high”

29. Common features • Do not consume themselves: no changes in quantity and quality before and after the reactions • Do not change the equilibrium points: only enhance the reaction rates. • Apply to the thermodynamically allowable reactions • Reduce the activation energy

30. Unique features • Enzyme-catalyzed reactions have very highcatalytic efficiency. • Enzyme have a high degree of specificity for their substrates. • Enzymatic activities are highly regulated in response to the external changes.

31. High Catalytic power: the ratio of E-catalyzed rate of a reaction to the uncatalyzed rate • Enzyme display enormous catalytic power, accelerating reaction rates as much as 1016 over uncatalyzed levels. Urease is a good example: Catalyzed rate: 3x104/sec Uncatalyzed rate: 3x10 -10 /sec Ratio is 1x1014 !

32. High Specificity: refers to the ability of an enzyme to discriminate between two competing substrates and catalyze one specific reaction • Specificity: the selective qualities of an enzyme (selecting substrate) • Based on structural complementarity, enzyme recognize substrate. • Unlike conventional catalysts, enzymes demonstrate the ability to distinguish different substrates.

33. Enzymes are highly specific both in the reaction catalyzed and in their choice of substrates. • An enzyme usually catalyzes a single chemical reaction or a set of closely related reactions. • There are three types of substrate specificities. • Absolute specificity • Relative specificity • Stereospecificity

34. Absolute specificity • Enzymes can recognize only one type of substrate and implement their catalytic functions • . For example, • urease only hydrolyzes urea to form NH3 and CO2. • not methylurea.

35. Relative specificity • Enzymes catalyze one class of substrates or one kind of chemical bond in the same type • Protein kinase A ,C,G, To phospharylate the –OH group of serine and threonine in the substrate proteins, leading to the activation of proteins.

36. stereospecificity • The enzyme can act on only one form of isomers of the substrates. • Lactate dehydrogenase can recognize only the L-form but the D-form lactate.

37. High regulation • Enzyme-catalyzed reactions can be regulated in response to the external stimuli, satisfying the needs of biological processes • Regulation can be accomplished through varying the enzyme quantity, adjusting the enzymatic activity, or changing the substrate concentration.

38. Classification of enzyme • 1956, International Commission on Enzymes (IUBMB) • -Classified by the types of reactions that they catalyze • 6 classes recognize Oxidoreductases, Transferases, Hydrolases, Lyases, Isomerases,Ligases • With each class subdivided into further subclasses

40. (1) Oxidoreductase • Catalyze oxidation-reduction reactions. • catalyze the transfer of hydrogen atoms and electrons • Contains dehydrogenase and oxidase. • For example:lactate dehydrogenase, peroxidase.

41. (2) Transferase • Catalyze transfers of groups between donors and acceptors. • For example:glutamate-pyruvate trasaminase (GPT), acyltransferase.

42. (3) Hydrolase • Catalyze cleavage of bonds by addition of water. • For example:pyrophosphatase, peptidase.

43. (4) Lyase • Catalyze lysis of a substrate, generating a double bond or adding a substrate to a double bond of a second substrate. • For example:pyruvate decarboxylase, aldolase.

44. (5) Isomerase • Catalyze racemization of optical or geometric isomers and certain intramolecular oxidation-reduction reactions. • For example:alanine racemase, mutase.

45. (6) Ligase or Synthetase • Join two molecules at the expense of a high energy phosphate bond of ATP. • For example:glutamine synthetase, carboxylase.

47. Enzyme nomenclature • Traditional nomenclature: • the suffix –ase :urease, phosphatase, • some bearing little resemblance to their activity: trypsin and pepsin (proteases) — easy confuse • Systematic nomenclature: • Each enzyme is given a systematic name • which identifies the reaction catalyzed. • Hexokinase----ATP: D-hexose-6- • phosphotrasferase

48. A four digital number can precisely identify all enzymes • Each enzyme is assigned a four-digit number with the first digit denoting the class it belongs, the other three further clarifications on the reaction catalyzed. Hexokinase: E.C. 2.7.1.1 • class 2, transferase

49. Systematic name Substrates are stated first, followed by the reaction type to which the ending-ase is affixed. Serial number: EC. X.X.X.X （ EC——Enzymes Commission ） Lactate:NAD+ oxidoreductase EC1.1.1. 27 Major class----oxidoreductase Subclass----oxidation group CHOH Subsubclass----NAD+ as hydrogen acceptor No. in this subsubclass Common name:Lactate dehydrogenase (LDH)

50. Some definitions • Apoenzyme = the protein part of an enzyme without coenzymes or prosthetic groups that are required for the enzyme to have activity. (Note: many enzymes do not have coenzymes or prosthetic groups bound to them). • Coenzyme = small organic or inorganic molecules which are bound to the apoenzyme and are required for the enzyme to catalyze the chemical reaction. • Prosthetic group = similar to a coenzyme, but is tightly bound to the apoenzyme. Heme is a prosthetic group in cytochrome c and hemoglobin. • Holoenzyme = the apoenzyme with the coenzyme or prosthetic group bound to it (i.e. the active form of the enzyme).