Introduction to Enzymes Lysozyme active site Chapter 8 Part 1 HIV-1 Protease with bound Inhibitor Dr. Ray
How Enzymes Function What structural features allow an enzyme to have its unique biochemical function? In Chapter 8 we will examine what enzymes are, how the rates of enzyme catalyzed reactions are studied (kinetics), and the effects that different types of inhibitors have on slowing down the rate of enzyme catalyzed reactions. In Chapter 9 we will study the mechanism of Serine Proteases like Trypsin and Chymotrypsin, how they hydrolize peptide bonds (catalysis) and how they can cut after particular residues (specificity). In Chapter 10 we will examine how enzymes are regulated. In Chapter 7 we have already examined: - the structural features (3 o & 4 o ) of the Mb and Hb active sites that allow the function of these proteins in reversible O 2 binding and transport - structural features of the Hb tetramer that allows it to cooperatively bind O 2 with different affinities in different environments within an organism - How small molecules fine tune the reactivity of Hb
Key Concept Map for Enzymes: Properties, Kinetics Inhibition Kinetic Parameters: K M substrate binding affinity k cat the turnover number k cat / K M catalytic efficiency V max maximal velocity V max = k cat [E] tot V o = V max [S] --------- K M + [S] 1 = K M 1 + 1 ---- ------- ---- ------ V o V max [S] V max Ref: Lippincott's Illustrated Reviews: Biochemistry, 3 rd Ed., Fig 5.23
Enzymes: The Catalysts of Biological Systems 1.Enzymes (E) are proteins capable of catalyzing chemical reactions, and speeding up the rates of biochemical reactions. 2.Proteins are highly effective in catalyzing chemical reactions, because of their capacity of binding specifically a wide range of molecules. 3.The main two features of enzymes are: catalytic power (rate enhancement: k cat /k uncat = 10 6 10 17 ) high substrate specificity (ability to discriminate between two competing substrates) 4.Enzyme s utilize the full repertoire of intermolecular forces to bring substrates together in an optimal orientation to make or break chemical bonds: Q: What properties surround substrates with protein so reaction distinguish enzyme s occurs in a very controlled environment from other catalysts? use organic and inorganic cofactors 5.Most enzymes are proteins, though some ribozymes exist (catalytically active RNA molecules).
Enzymes Active Sites 1. The active site of an enzyme is the region where substrates bind (prosthetic groups bind) 2. Active sites take up a relatively small part of the total volume of an enzyme. 3. Active sites are 3-D entities formed by groups that come from different parts of the linear amino acid sequence. 4. Active sites are clefts or crevices, rich in nonpolar (hydrophobic) residues and in which water is excluded (unless its a reactant). 5. The active site residues that participate in the making and breaking of bonds are called catalytic groups, and usually have polar or charged sidechains serving specific functions in the catalytic mechanism. 6. The specificity of binding substrate depends on the defined 3D arrangement of amino acid residues in the active site. Some active sites residues participate in catalysis while others participate in substrate binding via many non-covalent interactions. Lysozyme: active site residues shown in color
HIV Protease with bound Inhibitor Enzymes Active Sites Thermas aquaticus (Taq) RNA Polymerase http://bcs.wiley.com/hebcs/books?action=mininav&bcsid=2350&itemid= 0471214957&assetId=70681&resourceId=5828 DNA = blue/green RNA = red Protein = white http://cbm.msoe.edu/modgallery/rnapol/rnapolviewer.swf
Classes of Enzymes Know 6 main classes and be able to categorize specific enzymes Enzyme Nomenclature (name ends in ase) Classify enzymes by reaction type. Most enzymes catalyze the transfer of electrons, atoms or functional groups. 1. Oxidoreductases (oxidation-reduction reactions), e.g. lactate dehydrogenase 2. Transferases (group transfer reactions), e.g. protein kinase A 3. Hydrolases (hydrolysis reactions), e.g. chymotrypsin 4. Lyases (addition or removal of groups to form double bonds), e.g. fumarase 5. Isomerases (isomerization reactions), e.g. triose phosphate isomerase 6. Ligases (ligation of two substrates at the expense of ATP hydrolysis), e.g. aminoacyl-trna-synthetase Each enzyme is precisely defined by four numbers, which specify the class (type of transfer reaction), the type of donor (substrate), the type of acceptor, and the enzyme name, for example: E.C.1.1.3.17 Oxidoreductases Choline oxidase Acting on the CH-OH group of donors With oxygen as acceptor For the official Enzyme Nomenclature, see: http://us.expasy.org/enzyme/
International Classification of Enzymes Based on Type of Reaction Catalyzed Main Class Type of Reaction Catalyzed Subclasses Type of Reaction Catalyzed 1 Oxido- reductases Transfer of Electrons (oxidation-reduction) Dehydrogenases Oxidases Reductases Introduction of a Double Bond by Removal of H 2 Oxidation Reduction 2 Transferases Transfer of Functional Groups 3 Hydrolases Hydrolysis Reactions (Group transfer to H 2 O) 4 Lyases Group Addition / Elimination to Form Double Bonds 5 Isomerases Isomerization (Intramolecular Group Transfers) 6 Ligases Bond Formation Coupled to ATP Hydrolysis Kinases Transaminases Proteases Nucleases Lipases Glycosylases Decarboxylases Dehydrases Epimerases Carboxylases Synthetases Transfer a Phosphate Group Transfer a Amino Group Hydrolysis of a Amide Group Hydrolysis of a Phosphate Group Hydrolysis of an Ester Group Hydrolysis of an Acetal Group Loss of CO 2 Loss of H 2 O Isomerization of a Stereogenic Center Addition of CO 2 Formation of a New Bond
What class of enzyme will catalyze the following reactions? (Choose among the six major classes of enzymes) (1) (4) (2) (5) (3) 1- Amide: 2- Ester: 3- Phosphate transfer: 4- Pyruvate Lactate: 5- DHAP - GAP conversion: 6- Carbonic Anhydrase reaction: (6)
Enzyme Catalysis 1) What structural features at the active site of an enzyme allow it to have its unique biological function? use many catalytic mechanisms together to achieve high rate enhancements bring multiple functional groups and Facilitate acid-base reactions substrates together React with substrates as: enzyme is flexible, so can adjust Nucleophile or Electrophile to bound ligand H-bond partner has critically placed functional groups in active site that interact favorably with bound substrate and lower the energy of the highest energy point in the reaction: 1. acids.. 2. bases 3. polar groups 4. charged groups 5. metal cations Give examples of each type of functional group in proteins What functions do they serve?
Rates of Enzyme Catalyzed Reactions 1) Why are the rates of biochemical reactions important? Biomolecules exist in a dynamic steady state, where their rate of formation is matched by their rate of breakdown: 2) How much effect do enzymes have on the rates of reactions? Cells eliminate harmful hydrogen peroxide by: 2 H 2 O 2 2 H 2 O + O 2 Ref: Campbell and Farrell, Biochemistry, 4 th Ed., Table 5.1 3) Why are proteins stable in water, since peptide bond hydrolysis is a spontaneous reaction? the reaction is but the reaction is, so need a catalyst to: - speed up the reaction - control when reaction occurs
Free Energy Changes are used to Understand Enzyme Function Free energy change of a reaction (DG) tells us if a reaction can occur spontaneously. DG = DH - TDS Free Energy Enthalpy Entropy Reactions can be driven by: enthalpy (heat of reaction) or entropy (disorder) changes, or both. Figure 1.14. Protein Folding. Protein folding entails the transition from a disordered mixture of unfolded molecules to a relatively uniform solution of folded protein molecules. For now SKIP details of text section DG of a reaction is related to Keq. We will study this later with metabolism (unit 3). Figure 1.15. The Hydrophobic Effect. The aggregation of nonpolar groups in water leads to an increase in entropy owing to the release of water molecules into bulk water.
Free Energy Changes and Equilibrium A + B C + D DG = DH - TDS The DG of a reaction tells us whether a reaction can or cannot occur spontaneously, or whether the system is at equilibrium. 1. DG < 0 reaction occur spontaneously from A, B C, D This is an exergonic reaction. 2. DG > 0 reaction occur spontaneously from C, D A, B An input of free energy is needed to drive this reaction, which is endergonic. 3. DG = 0 reaction is at equilibrium There is no net change in concentrations of all species. 4. The DG of a reaction is independent of the path (molecular mechanism) of the transformation. So the free energy change (the driving force) for an enzyme catalyzed reaction and an uncatalyzed one is the same. 5. DG provides no information about the rate of a reaction!
Equilibrium Thermodynamics and Kinetics The standard free energy change of reaction (DG ): is the energy difference between the products & reactants is related to the equilibrium constant (K eq ) of a reaction: For reaction: A + B C + D Ex: Amide + H 2 O Amine + Carboxylate Thermodynamics (Chapter 15) allows us to answer the question: Q: Is the reaction spontaneous? Will it proceed as written? Kinetics allows us to answer the question: Q: How fast is the reaction? o' K eq' C DG RT log K e D A B ' eq Activation Energy Spontaneous does NOT mean instantaneous. Spontaneous means if a reaction can occur (can overcome the activation energy), the products are energetically more stable than the reactants. Reaction rates are linked to Activation Energy ( DG ), the energy required to initiate the conversion of reactants to products. DG -
Catalytic Power of Enzymes Enzyme s accelerate reactions by factors of at least a million times. Most of the reactions in biological systems would NOT OCCUR at any perceptible rate in the absence of enzymes. Rate enhancement = Enzyme Catalyzed Rate = 10 6-10 17 Uncatalyzed Rate of Nonenzymatic Reaction Rate Enhancement by = 578 = 1.9 x 10 11 Carboxypeptidase A 3.0 x 10-9 Preferential Transition state binding (11.15) http://higheredbcs.wiley.com/legacy/college/voet/0471214957/animated_figures/ch11/f11-15.html
Specificity of Enzymes 1. Enzyme s are highly specific in the reaction catalyzed and the choice of their substrates (S), so enzymes can discriminate between competing substrates Specificity arises from precise interaction (mostly noncovalent) of the substrate with the enzyme. This precision is a result of the intricate 3D structure of the enzyme. (Protein Structure Function) 2. An Enzyme usually catalyzes a single reaction or a set of closely related reactions. Trypsin catalyzes the hydrolysis of peptide bonds on the C-side of Arg and Lys residues, during digestion. Thrombin catalyzes the hydrolysis of peptide bonds between Arg and Gly in particular sequences only important in blood clotting. 3. Enzymes are stereospecific Oxidation of Ethanol by Yeast Alcohol Dehydrogenase (YADH) 1) Which reactant is oxidized and which is reduced? CH 3 CH 2 OH is NAD + is 2) Examples belong to which enzyme class? Trypsin - DH- = deuterated form of NADH
Review of Chemical Kinetics (see extra Review PowerPoint notes read at home) For the reaction: aa + bb cc + dd Rate law equation describes how reaction velocity (D[product] with time) depends on concentrations of reactant and products: Rate = k [A] [B] V o = -d[a]/dt The proportionality constant k is the rate constant V o is initial reaction rate (change in concentration of reactants per unit time) First order reactions: Second order reactions: 1) How do enzyme s alter the rate of a reaction? 2) Do they affect the the rate constant k or the concentrations of substrates? Rate = k [ A ] Rate = k [ A ] 2
Rate Laws and Reaction Mechanism Consider SN2 reaction: CH 3 Cl + OH - CH 3 OH + Cl - The nucleophile OH - attacks the electrophile CH 3 Cl from the back The resulting transient species with both OH - and Cl - simultaneously bound to C is the high energy transition state (activated complex) C-Cl bond breaks and the C-OH bond is completed This is called: Substitution, Nucleophilic, Bimolecular ( SN2 ) with a rate law: Rate = [CH 3 Cl][OH - ] for overall rxn determined from expt *mechanism depends on concentrations of BOTH reactants Consider SN1 reaction: (CH 3 ) 3 CCl + OH - (CH 3 ) 3 COH + Cl - This is called: Substitution, Nucleophilic, Unimolecular ( SN1 ) with a rate law: Rate = [(CH 3 ) 3 CCl] * mechanism only depends on concentration of 1) Draw the mechanism for SN1 and SN2 CARBOCATION reactions based on their rate laws. 2) What is the difference in the stereochemistry of the products of these two reactions?
Catalysis and Reaction Mechanism The balanced chemical equation provides information about the beginning and end of reaction. The reaction mechanism gives the path of the reaction, a very detailed picture of which bonds are broken and formed during the course of a reaction. Individual steps at the molecular level (including the movement of electron pairs), are described. Elementary step is any process that occurs in a single step in the mechanism. Elementary steps must add to give the balanced chemical equation. Intermediate is a species which is produced in an elementary step, then used in a later elementary step, but which is not a reactant or product. (intermediate has some stability, it is not fleeting like the transition state) Catalyst is present in very small quantities relative to reactants. A catalyst is a species that is used in elementary step, and regenerated in a later step in the reaction pathway. It is not a reactant or product. It changes the rate of a chemical reaction and makes the reaction go faster.
Rate Laws of Elementary Steps Molecularity: the number of reactant molecules Examples: present in an elementary step k Unimolecular: one molecule in the elementary step A B Bimolecular: two molecules in the elementary step A + B C Termolecular: three molecules in the elementary step (termolecular processes are not common, since they are statistically improbable) A + B + C D The rate law of an elementary step is determined by its molecularity: Unimolecular processes are first order Rate = k [A] Bimolecular processes are second order Rate = k [A][B] Termolecular processes are third order Rate = k [A][B][C] 1) If a mechanistic step in a reaction is known to be an elementary step, what is the rate law for this step? - this is a molecular rxn k 2 ES P + E Rate =
Enzyme Catalyzed Reactions E = enzyme S = substrate P = product I = inhibitor Uncatalyzed reaction: S S P Enzyme catalyzed reaction: ES E + S E. S E + P E. S = enzyme-substrate complex k uncat = rate constant for reaction in a testube in the lab k cat = maximal rate constant in the presence of enzyme, is rate constant for the rate determining (slowest, RDS) step in the mechanism at saturating substrate concentrations Examine different classes of inhibitors, which are kinetically distinguishable E + I E. I no reaction 1. Irreversible (inactivators) 2. Reversible (competitive and noncompetitive)
The E-S Model for Catalysis by Enzymes 1. Substrates bind to active sites through multiple weak noncovalent interactions (hydrogen bonds, dipole-dipole interactions, saltbridges, van der Waals interactions, and hydrophobic interactions). Example: the enzyme ribonuclease (RNAse) forms hydrogen bonds with the uridine component of its RNA substrate. 2. Substrates fit snugly into the active site of an enzyme, since the 3D surface of the interacting species are complementary. 3. Reactions occur within an Enzyme Substrate (E-S) complex. Enzyme
Models for ES Complex Formation To fit in the active site, a substrate must have a matching shape, and complementary functional groups. Lock and key model (proposed by Emil Fisher in 1890). In this model, the active site of the unbound enzyme is preformed complementary in shape to the substrate. Induced fit model (proposed by Daniel E. Koshland in 1958). In this model, the enzyme changes shape upon substrate binding. The active site forms a shape complementary to the substrate only after the substrate has been bound weakly. http://www.wiley.com//college/boyer/0470003790/animations/enzyme_binding/ enzyme_binding.htm Binding Models
Models of Enzyme Ligand interactions: 1) Lock and Key Model: enzymes are structurally complementary to their substrates, so fit together like a key fitting into a lock both partners have correct geometries (and functional groups in correct orientations) prior to their interaction 2) Induced Fit Model: change in conformation of an enzyme in response to substrate binding, that renders the enzyme catalytically active Induced fit often places the catalytic groups in the right position for efficient catalysis! result is that shape of binding site of enzyme better conforms to the shape of the ligand (to achieve good geometric and electrostatic complementarity) enzymes are flexible, so small conformational adjustments can occur after initial loose enzyme-substrate association occurs, to form a tightly bound E. S complex
Substrate Induced Conformational changes in Hexokinase Induced Fit Model of Enzyme Ligand Interactions + Glucose (a) Open substrate-free form - glucose hexokinase 1. What type of reaction is catalyzed by a kinase? (b) Closed substrate-bound (purple glucose) form - flap folds down over substrate after it binds so that substrate is almost completely surrounded by the enzyme, and water is excluded. Class of Enzyme? reaction of phosphate from ATP to sugar hexose, without hydrolyzing ATP. Need to keep H 2 O out of active site when both substrates are bound, so enzyme completely surrounds the substrate.
Michaelis-Menten Model of Enzyme Kinetics ES E + S E.S E + P Michaelis-Menten model: Reversible binding and reaction of a single molecule of substrate within an enzyme active site, where a reactant is converted to product by a different mechanism than in solution. The product leaves active site and a new molecule of substrate binds to the Enzyme (thus the Enzyme turns over ). Much of the catalytic power of enzymes comes from bringing substrates together in favorable orientations in enzyme-substrate complexes (ES) in the active site. Most enzymes are highly selective in their binding of substrates.
Multistep Enzyme Catalyzed Reactions Consider a reaction mechanism consisting of more than one elementary step (a single step in a reaction mechanism): k 2 = k cat If k 2 is RDS: ES P + E then rate law for entire reaction is: Rate = k 2 [ES] Rate-determining step (RDS) in a multi-step mechanism is: the slowest of the elementary steps. The RDS is the bottleneck reaction in which species produced rapidly (in earlier steps) must wait to get through. Once thru the RDS, any remaining steps occur very quickly. Therefore, the rate-determining (RDS) step governs the OVERALL RATE LAW for the entire reaction. k 2
Reaction Kinetics Consider the reaction: S P Enzymes accelerate the rate of attainment of equilibria but do NOT shift their positions. Equilibrium position depends only on energy of products & reactants. 1) Draw an energy level diagram for this chemical reaction: 2) Which point in the reaction coordinate has the highest energy? The transition state (TS ) is the most unstable species, with the highest energy in the reaction coordinate. DG rxn = free energy of reaction, is the difference in free energy between the products and the reactants 3) Is the reaction shown exergonic or endergonic? 28
Reaction Kinetics The chemical reaction S P can now be written considering the role of the transition state (TS ): S TS P DG rxn In the TS covalent bonds are being formed and/or broken unstable The free-energy of activation (DG ) is the difference between the free energy of the substrate and that of the transition state, which is also called activation energy (E a ). 4) Can a reaction be very slow, even though it has a negative DG rxn? rate depends on, but equilibrium position depends on 5) How can the reaction be made faster? The rate of a chemical reaction is inversely proportional to the free energy of activation (DG ): the LOWER the free-energy of activation, the HIGHER the RATE of reaction!
Reaction Kinetics Enzyme catalysts lower the transition state energy of the reaction, thus decrease the activation energy: DDG = the reduction in DG by the catalyst DDG = DG (uncatalyzed) - DG (catalyzed) DDG shows the efficiency of the catalyst S TS P DDG (the reduction of DG by the catalyst) Enzyme catalysts accelerate the forward (S P) and reverse (P S) reactions, to the same extent. Example: Carbonic Anhydrase catalyzes H 2 O + CO 2 H 2 CO 3 reaction both ways in red blood cells: 6) Do enzymes alter reaction equilibria by changing the free energy of reactants and products, or the distribution of products? 30 alter thermodynamics
Enzyme Catalyzed Reactions Enzyme active site is most complementary to TS, not to substrate. 1) Do enzymes alter reaction rates by changing the mechanism of reaction (details of electron flow)?, the reaction occurs by mechanism The binding of S and E creates a new reaction pathway (new mechanism) whose TS has a lower energy. the activation barrier is now lower the rate of reaction is faster! (A) Uncataylzed reaction: S P (B) Enzyme cataylzed reaction: E + S ES E + P TS 2) Is there an intermediate species in plot B? 3) Where is the ES complex in plot B? Different reaction coordinate diagram reflects different mechanism from uncatalyzed reaction. In order to understand enzyme function, regulation and control, we need to first understand Enzyme Kinetics. 31
Hypothetical enzyme stickase 1) Which species binds most tightly to an enzyme catalyst (substrate, product, or something else)? Desired reaction (bend metal stick): No reaction, since S within E is more stable than by itself 2) Which species in the enzyme catalyzed reaction is the transition state? ES The essence of enzymatic catalysis is specific binding to the transition state. TS Very low energy ES DDG ES has same energy as S TS is stabilized by binding to enzyme, so faster reaction 32 since energy of TS is lowered
Enzyme Active Site Pocket is complementary to the structure of the TRANSITION STATE of a Reaction NOT to the structure of the substrate! hypothetical enzyme stickase Enzyme complementary to the TS will help to destabilize the substrate (will bend stick), resulting in catalysis of reaction 33 Ref: Lehninger Biochemistry
Enzymes, Activation Energy and Rate Constants 1. How do enzyme s alter the rate of a reaction? Rate = k [A] [B] Catalysts enhance reaction rate (speed of a reaction) by providing a lower energy route (different mechanism) between reactant & product! 2. Do enzymes affect the rate constant k or the concentrations of substrates? Most reaction-rate data obeys the Arrhenius Equation: So can experimentally study the temperature dependence of a reaction in order to find rate constant (k) values: k k is the rate constant E a is the activation energy = DG = energy difference between energy of reactants and energy of transition state R is the gas constant (8.314 J/K-mol) T is the temperature in Kelvin A is called the frequency factor, which is a measure of the probability of a favorable collision (between reactants) Ae E a RT 34 = E a
Enzymes, Activation Energy and Rate Constants rate = k[a][b] = E a k Ae E a RT By lowering the activation energy (E a = DG ) of a reaction: enzymes increase the value of the rate constant (k) of a reaction which increases the rate (speed) of the reaction, for the same concentration of substrates Enzymes increase rate constant (k), thus increase the rate of reaction!
Michaelis-Menten Kinetics ES Activation Energy fast Slow RDS The rate-determining (RDS) step governs the OVERALL RATE LAW for the entire reaction: ES complex Transition State ES Rate = k 2 [ES] E + P
1) Are catalytically active enzymes made up entirely of amino acids? The simple components (monomers) of the four classes of biomolecules: Amino acids Nucleotides Fatty acids Monosaccharides are precursors to many other kinds of biomolecules Components of Enzymes: Functional Roles of Biomolecules Holoenzyme = Cofactor + Apoenzyme (enzyme only has biological activity when both are present) Cofactor (coenzyme) small non-amino acid part of an enzyme Apoenzyme amino acid component of entire (holo) enzyme
Many Enzymes Require Cofactors Cofactor (or coenzyme) = small molecule that is required for the catalytic activity of an enzyme. Non-amino acid molecule can be organic molecule, metal ion, or organometallic complex. Coenzymes can be either: Cosubstrate = a small organic molecule that only transiently associates with an enzyme. (example: NAD +, FAD) Prosthetic Group = a molecule that is permanently and tightly bound to an enzyme (often, but not always covalently). (example: HEME) Ref: Voet, Voet & Pratt, Biochemistry Cofactor Enzyme This table is for Coenzyme A (CoA) Citrate Synthase reference, you do not need to know details Nicotinamide adenine dinucleotide Lactate dehydrogenase Tetrahydrofolate Thymidylate synthase (U T)
Many Enzymes Require Cofactors Holoenzyme = Cofactor + Apoenzyme Holoenzyme only has biological activity when cofactor is bound. Cofactors are often derived from dietary vitamins and minerals: Vitamins small organic molecules that must be obtained in the diet and are required in trace amounts for proper growth Structure of the vitamins nicotinamide and nicotinic acid, the redox active components of the coenzymes NAD + and NADP + that shuttle reducing equivalents throughout metabolism. These cofactors are cosubstrates for dehydrogenase enzymes, so associate transiently Once reduced, they transport electrons released by fuel oxidation (glucose metabolism) into the mitochondrial electron transport chain for ATP synthesis. NADH and FADH 2 are MOBILE electron carriers essential for metabolism!
Vitamins small organic molecules that must be obtained in the diet and are required in trace amounts for proper growth FYI Chemical Groups Transferred This Table lists the 13 known vitamins required in the human diet, their enzyme functions, symptoms in patients who have a deficiency of this vitamin, and the type of chemical group transferred. Aldehydes Electrons Amino Groups Hydride Ion ( :H - ) One Carbon Groups H-atoms, Alkyl Groups Acyl Groups CO 2 This table is for reference, you do not need to know details
Many Enzymes Require Cofactors (Table 8.2) Inorganic ions are needed in metalloproteins and metalloenzymes as inorganic cofactors. They are supplied by foods or vitamins and minerals. Cofactor Metal: Zn 2+ Zn 2+ Mg 2+ Mg 2+ Ni 2+ Mo Se Mn 2+ K + FYI This table is for reference, you do not need to know details Fe 2+ /Fe 3+ and Cu + /Cu 2+ Enzyme Cytochrome c electron transfer Carbonic anhydrase His Carboxypeptidase EcoRV Hexokinase Urease Hemoglobin Nitrate reductase O 2 transport Glutathione peroxidase Superoxide dismutase Propionyl CoA carboxylase Redox enzymes His Cofactors such as hemes (Fe-porphyrins) are modular units and participate in many different kinds of reactions, by varying active site covalent interactions (axial ligands) and noncovalent interactions. Met
Summary of Kinetic Parameters and Equations Association/Dissociation Fast Michealis constant (K M ) Product Formation Slow V o = V max [S] K M + [S] 1 = K M 1 + 1 ---- ------- ---- ------ V o V max [S] V max Know Kinetic Parameters: V max = k cat [E] tot = k 2 [ES] is the maximal velocity when the enzyme active site is saturated with substrate K M Substrate Binding Affinity (when k -1 >> k 2 ): K M = k -1 / k 1 k cat is the turnover number: k cat = K M how active E is at catalyzing rxn how tightly E binds S k cat = V max [E] T k cat / K M is a measure of enzyme s catalytic efficiency when limited amount of substrate is available [S] << K M The M-M model holds when: (1) at initial stage of reaction, (2) have steady state conditions, and (3) [S] >> [E] http://higheredbcs.wiley.com/legacy/college/voet/0471214 957/guided_ex/michaelis_menten/michaelis_menten.html