Chapter 6: Enzymes

Question 1

cofactor

Answer 1

• reusable non-protein molecules that doesn’t contain carbon (inorganic)
• Usually are metal ions such as iron, zinc, cobalt, and copper that loosely bind to an enzyme’s active site
• a substance that increases the rate of a chemical reaction
• can be considered “helper molecules” that assist in
• They must also be supplemented in the diet as most organisms do not naturally synthesize metal ions.

Question 2

coenzyme

Answer 2

• an organic non-protein compound that binds with an enzyme to catalyze a reaction
• often broadly called cofactors, but they are chemically different
• it cannot function alone, but can be reused several times when paired with an enzyme

Question 3

prosthetic groups

Answer 3

• organic vitamins, sugars, lipids, or inorganic metal ions
• unlike coenzymes or cofactors, these groups bind very tightly or covalently to an enzyme to aid in catalyzing reactions
• often used in cellular respiration and photosynthesis.

Question 4

A complete, catalytically active enzyme together with its bound coenzyme and/or metal ions is called a _____. The protein part of such an enzyme is called the _____ or _____

Answer 4

• holoenzyme
• apoenzyme
• apoprotein

Question 5

active site

Answer 5

• region of an enzyme that binds substrate molecules
• This is crucial for the enzyme’s catalytic activity.

Question 6

substrate, S

Answer 6

The molecule that is bound in the active site and acted upon by the enzyme

Question 7

The surface of the active site is lined with _____ _____ _____ with substituent groups that bind the substrate and catalyze its chemical transformation. Often, the active site _____ a substrate, sequestering it completely from solution.

Answer 7

• amino acid residues
• encloses

Question 8

Catalysts do not affect reaction _____

Answer 8

equilibria

Question 9

energy in biological systems is described in terms of

Answer 9

free energy, G.

Question 10

ground state

Answer 10

• starting point for either the forward or the reverse reaction
• the contribution to the free energy of the system by an average molecule (S or P) under a given set of condition

Question 11

exothermic reactions

Answer 11

• the system loses heat as the surroundings warm up
• heat energy is being released from the system to the surroundings
• -ΔH = NEGATIVE

Question 12

endothermic reactions

Answer 12

• the system gains heat as the surroundings cool down
• heat energy is being absorbed by the system from the surroundings
• ΔH = POSITIVE

Question 13

exothermic reaction coodinate

Answer 13

• y axis = potential energy
• x axis = reaction pathway
• forward reaction: reactants on left, products on right
• reverse reaction: flip side reactants & products are on
• activated complex: is an intermediate compound w/higher energy than both reactants and products
• ΔH represents the difference between enthalpy of reactants and products
  • ΔH = H_PRODUCTS – H_REACTANTS
• _​The step with the highest activation energy (ΔG^‡) is the slowest step reaction
• The step with the lowest activation energy (ΔG^‡) is the fastest step in the reaction
• The reaction cannot proceed faster than the rate of the slowest elementary step

Question 14

endothermic reaction coodinate

Answer 14

• y axis = potential energy
• x axis = reaction pathway
• activated complex: is an intermediate compound w/higher energy than both reactants and products
• ΔH represents the difference between enthalpy of reactants and products
  • ΔH = H_PRODUCTS – H_REACTANTS
• _​The step with the highest activation energy (ΔG^‡) is the slowest step reaction
• The step with the lowest activation energy (ΔG^‡) is the fastest step in the reaction
• The reaction cannot proceed faster than the rate of the slowest elementary step

Question 15

reaction coordinate

Answer 15

• A: potential energy of reactants
• B: ΔG^‡: activation energy: energy needed to start reaction
• C: ΔG^‡: activation energy: reverse reaction
• D: ΔH: energy of reaction
• E: potential energy of products

Question 16

Answer 16

Question 17

ΔG^o, the standard free-energy change

Answer 17

• describes the free-energy changes for reactions
• a standard set of conditions
  
  • temperature 298 K
  • partial pressure of each gas 1 atm, or 101.3 kPa
  • concentration of each solute 1 M

Question 18

ΔG’^o, the biochemical standard free-energy change

Answer 18

• used because biochemical systems commonly involve H⁺ concentrations far below 1 M
• it is the standard free-energy change at pH 7.0

Question 19

reaction coordinate diagram

Answer 19

• S = Substrate, P = Product
• free energy of the system is plotted against the progress of the reaction S → P
• description of the energy changes during the reaction
• horizontal axis (reaction coordinate) reflects the progressive chemical changes (e.g., bond breakage or formation) as S is converted to P
• activation energies, ΔG^‡, for the S → P and P → S reactions are indicated
• ΔG’^o
  
  • standard free-energy change in the direction S → P
  • exergonic: it’s negative, the free energy of the ground state of P is lower than that of S
  • at equilibrium there is more P than S (the equilibrium favors P)

Question 20

transition state

Answer 20

• The rate of a reaction is dependent on an energy barrier between reactants and products
  
  • energy required for alignment of reacting groups, formation of transient unstable charges, bond rearrangements, and other transformations required for the reaction to proceed in either direction
• depicted by the energy “hill” in reaction coodinate
• molecules must overcome this barrier with a higher energy level
• the top of the energy hill represents a moment where things could go either way, decay to either substrate or product is equally likely
• not a chemical species with any significant stability and should not be confused with a reaction intermediate

Question 21

activation energy, ΔG^‡

Answer 21

• difference between the energy levels of the ground state and the transition state
• rate of a reaction reflects ΔG^‡
• a higher ΔG^‡ corresponds to a slower reaction
• Reaction rates can be increased by raising the temperature and/or pressure, thereby increasing the number of molecules with sufficient energy to overcome the energy barrier
• ΔG^‡ can be lowered by adding a catalyst

Question 22

reaction intermediates

Answer 22

• A reaction intermediate is any species on the reaction pathway that has a finite chemical lifetime (longer than a molecular vibration)
• occupy valleys in the reaction coordinate diagram

Question 23

rate-limiting step

Answer 23

• the step (or steps) with the highest activation energy determines the overall rate
• it’s the highest-energy point in the reaction coordiante diagram
• can vary with reaction conditions, and for many enzymes several steps may have similar activation energies, which means they are all partially rate-limiting

Question 24

• Reaction equilibria is linked to the _____ _____-_____ for the reaction
• reaction rates are linked to the _____ _____

Answer 24

• standard free-energy change ΔG’º
• activation energy ΔG^‡

Question 25

equilibrium constant, K'_eq or K'

Answer 25

* denotes equilibrium: (S)ubstrate ⇔ (P)roducts * K'_eq = P / S

Question 26

for thermodynamics the relationship between K'_eq and ΔG'º can be described by the following formula

Answer 26

* ΔG'º = -RT ln K'_eq * R: gas constant, 8.315 J/mol • K * T: absolute temperature, 298 K (25 8C) * equilibrium constant is directly related to the standard free-energy change for the reaction * A large negative value for ΔG'º reflects a favorable reaction equilibrium * more product than substrate at equilibrium * doesn't mean the reaction will proceed at a rapid rate

Question 27

rate law or differential rate law

Answer 27

* rate law * expresses the relationship between the rate of the reaction and the concentration of the reactant * no products are involved * differential rate law * used to describe what is occurring on a molecular level during a reaction, whereas integrated rate laws are used for determining the reaction order and the value of the rate constant from experimental measurements

Question 28

rate law formula

Answer 28

* V = k[S]ⁿ * S: concentration of substrate (reactant) * k: rate constant * V: rate, velocity of the reaction. the amount of R that react per unit of time * n: reaction order

Question 29

rate constant (k)

Answer 29

* constant of proportionality * changes only with change in temp * Unit of k depends on order of reaction

Question 30

reaction order (n)

Answer 30

* usually integer can be fraction * determines how rate depends on [] of reactant * can only b determined experimentally

Question 31

overall order

Answer 31

the sum of the exponents for reactions with more than one reactant

Question 32

reaction order values: zero order n = 0 rate = k[S]⁰

Answer 32

* S = substrate = reactant, P = product * rate is independent of concentration of S * because [S]⁰ = 1 so rate is equal to k * concentration ↓ linearly * slope of line is constant → constant rate * occurs when amt of reactant available for reaction unaffected by changes to ttl amt of reactant * units of Ms^-1

Question 33

reaction order values: first order n = 1 rate = k[S]¹

Answer 33

* S = substrate = reactant, P = product * rate directly proportional to [S] * rate ↓ as reaction proceeds cuz [S] ↓ * slope of curve (rate) less steep w/time * If a first-order reaction has a rate constant k of 0.03 s^-1 this means that 3% of the available S will be converted to P in 1 s * units of s^-1

Question 34

reaction order values: second order n = 2 rate = k[S]² or rate = k[S₁][S₂]

Answer 34

* S = substrate = reactant, P = product * If a reaction rate depends on the concentration of two different compounds * rate proportional to square of [S] if the same compounds: rate = k[S]² * or proportional to the product of the two [S]: rate = k[S₁][S₂] * rate more sensitive to [S] * rate proportional to [S]² * slope flattens quicker than 1st order * initial rate ↑ 4x when [S] doubles

Question 35

rate / differential law concentration vs time graph

Answer 35

Question 36

rate / differential law rate vs concentration graph

Answer 36

Question 37

unit of k (rate constant) pattern

Answer 37

Question 38

From transition-state theory we can derive an expression that relates the magnitude of a rate constant to the activation energy:

Answer 38

* k: Boltzmann constant = 1.38064852 × 10^-23 m² kg s^-2 K^-1 * h: Planck’s constant = 6.62607015 × 10−34 joule second * The important point here is that the relationship between the rate constant k and the activation energy ΔG^‡ is inverse and exponential

Question 39

What are two sources of energy that lower the activation energy

Answer 39

Covalent and noncovalent interactions between enzymes and substrate

Question 40

What are two sources of energy that lower the activation energy? Covalent interactions between enzymes and substrates

Answer 40

* the rearrangement of covalent bonds during an enzyme-catalyzed reaction. * Catalytic functional groups on an enzyme may form a transient covalent bond with a substrate and activate it for reaction * or a group may be transiently transferred from the substrate to the enzyme * In many cases, these reactions occur only in the enzyme active site * these covalent interactions between enzymes and substrates lower the activation energy (and thereby accelerate the reaction) by providing an alternative, lower-energy reaction path.

Question 41

What are two sources of energy that lower the activation energy? noncovalent interactions

Answer 41

* critical to the formation of complexes between proteins and small molecules, including enzyme substrates * Much of the energy required to lower activation energies is derived from weak, noncovalent interactions between substrate and enzyme * interaction between substrate and enzyme in this complex is mediated by the same forces that stabilize protein structure, including hydrogen bonds and hydrophobic and ionic interactions * Formation of each weak interaction in the ES complex is accompanied by release of a small amount of free energy that stabilizes the interaction called the binding energy, ΔG_B

Question 42

binding energy, ΔG_B

Answer 42

* energy derived from enzyme-substrate interaction * stabilizes the enzyme-substrate complex * a major source of free energy used by enzymes to lower the activation energies of reactions * contributes to specificity as well as to catalysis * Weak interactions are optimized in the reaction transition state * enzyme active sites are complementary not to the substrates per se but to the transition states through which substrates pass as they are converted to products

Question 43

An enzyme completely complementary to its substrate would be a very _____ enzyme

Answer 43

poor

Question 44

optimal interactions between substrate and enzyme occur only in the _____ \_\_\_\_\_

Answer 44

transition state

Question 45

Some _____ interactions are formed in the enzyme substrate complex, but the full complement of such interactions is formed only when the substrate reaches the transition state. The _____ \_\_\_\_\_ released by the formation of these interactions partially offsets the energy required to reach the top of the energy hill. The summation of the unfavorable (positive) _____ \_\_\_\_\_ and the favorable (negative) _____ \_\_\_\_\_\_ results in a lower net activation energy

Answer 45

* weak * free energy (binding energy) * activation energy ΔG^‡ * binding energy ΔG_B

Question 46

weak binding interactions between the enzyme and the substrate provide a substantial driving force for _____ \_\_\_\_\_

Answer 46

enzymatic catalysis

Question 47

The weak interactions formed only in the transition state are those that make the _____ contribution to catalysis

Answer 47

primary

Question 48

requirement for multiple weak interactions to drive catalysis is one reason why enzymes (and some coenzymes) are so \_\_\_\_\_. An enzyme must provide _____ \_\_\_\_\_ for ionic, hydrogen-bond, and other interactions, and also must precisely _____ these groups so that binding energy is optimized in the transition state

Answer 48

* large * functional groups * position

Question 49

* There are 4 prominent physical and thermodynamic factors contributing to ΔG^‡, the barrier to reaction: * ____ \_\_\_\_\_ can be used to overcome all these barriers.

Answer 49

* 4 prominent physical and thermodynamic factors * the entropy (freedom of motion) of molecules in solution, reduces the possibility that they will react together * The solvation shell of hydrogenbonded water surrounds and helps to stabilize most biomolecules in aqueous solution * distortion of substrates that must occur in many reactions * Need for proper alignment of catalytic functional groups on the enzyme * Binding energy

Question 50

entropy reduction

Answer 50

* the large restriction in the motions of two substrates that are going to react * obvious benefit of binding substrates to an enzyme. * Binding energy holds the substrates in the proper orientation to react

Question 51

Studies have shown that constraining the _____ of two reactants can produce rate enhancements of many orders of magnitude

Answer 51

motion

Question 52

desolvation

Answer 52

* Enzyme-substrate interactions replace most or all of the hydrogen bonds between the substrate and water that would otherwise impede reaction * binding energy involving weak interactions formed only in the reaction transition state helps to compensate thermodynamically for any distortion, primarily electron redistribution, that the substrate must undergo to react

Question 53

induced fit

Answer 53

* enzyme itself usually undergoes a change in conformation when the substrate binds, induced by multiple weak interactions with the substrate * a network of coupled motions occurs throughout the enzyme that ultimately brings about the required changes in the active site * brings specific functional groups on the enzyme into the proper position to catalyze the reaction * conformational change also permits formation of additional weak bonding interactions in the transition state * new enzyme conformation has enhanced catalytic properties * a common feature of the reversible binding of ligands to proteins

Question 54

Once a substrate is bound to an enzyme, properly positioned catalytic _____ \_\_\_\_\_ aid in the cleavage and formation of bonds by a variety of mechanisms, including .... It generally involve transient _____ interaction with a substrate or group transfer to or from a substrate

Answer 54

* functional groups * general acid-base catalysis, covalent catalysis, and metal ion catalysis * covalent

Question 55

General Acid-Base Catalysis

Answer 55

* proton transfer * single most common reaction * occurs in most reactions that take place in cells * Many biochemical reactions involve the formation of unstable charged intermediates that break down rapidly to their constituent reactant impeding the reaction * Charged intermediates can often be stabilized by the transfer of protons to or from the substrate or intermediate to form a species that breaks down more readily to products * studied using nonenzymatic model reactions * can involve either the constituents of water alone or other weak proton donors or acceptors

Question 56

General Acid-Base Catalysis specific acid-base catalysis

Answer 56

* Catalysis of the type that uses only the H⁺ (H₃O⁺) or OH^- ions present in water * If protons are transferred between the intermediate and water faster than the intermediate breaks down to reactants, the intermediate is effectively stabilized every time it forms.

Question 57

general acid-base catalysis

Answer 57

* In many cases, however, water is not enough * refers to proton transfers mediated by weak acids and bases other than water * For nonenzymatic reactions in aqueous solutions * Many weak organic acids can supplement water as proton donors in this situation * weak organic bases can serve as proton acceptors.

Question 58

In the active site of an enzyme, where _____ may not be available as a proton donor or acceptor, general acid-base catalysis becomes crucial. A number of _____ \_\_\_\_\_ _____ \_\_\_\_\_ can and do take on the role of proton donors and acceptors. These groups can be precisely positioned in an enzyme active site to allow proton transfers, providing rate enhancements of the order of 10² to 10⁵. This type of catalysis occurs on the vast majority of \_\_\_\_\_

Answer 58

* water * amino acid side chains * enzymes

Question 59

Covalent Catalysis

Answer 59

* a transient covalent bond is formed between the enzyme and the substrate * results in catalysis only when the new pathway has a lower activation energy than the uncatalyzed pathway * A number of amino acid side chains can serve as nucleophiles in the formation of covalent bonds with substrates * always undergo further reaction to regenerate the free enzyme * covalent bond formed between the enzyme and the substrate can activate a substrate for further reaction in a manner that is usually specific to the particular group or coenzyme

Question 60

Metal Ion Catalysis

Answer 60

* Metals can participate in catalysis in several ways * Ionic interactions between an enzyme-bound metal and a substrate can help orient the substrate for reaction or stabilize charged reaction transition states

Question 61

Metal Ion Catalysis * Metals can also mediate \_\_\_\_\_-\_\_\_\_\_reactions by reversible changes in the metal ion’s oxidation state * Nearly a third of all known enzymes require one or more\_\_\_\_\_ _____ for catalytic activity.

Answer 61

* oxidation-reduction * metal ions

Question 62

Most enzymes combine several _____ strategies to bring about a rate enhancement

Answer 62

catalytic

Question 63

enzyme kinetics

Answer 63

* oldest approach to understanding enzyme mechanisms * one that remains most important * determines the rate of a reaction and how it changes in response to changes in experimental parameters

Question 64

enzyme kinetics - variables * E * S * P * [E_t] * k₁ * k-₁ * K₂ * k_-2 * [E_t]

Answer 64

* E = enzyme, * S = substrate * P = product * [E_t] = total enzyme, bound/unbound * k₁ = rate constant for binding of E to S forming the ES complex * k-₁ = rate constant for dissociation of ES to free E and S (reactants) * K₂ = * rate constant for catalytic rate * catalysis reaction producing the final reaction product and regenerating the free enzyme * rate limiting step. * k-₂ = rate constant for the reverse reaction of catalysis

Question 65

[E_t]

Answer 65

* total enzyme concentration * the sum of free/unbound enzyme [E] and substrate-bound enzyme [ES] * [E_t] = [E] + [ES] * Free or unbound enzyme [E] can then be represented by * [E] = [Et] - [ES] * Because [S] is ordinarily far greater than [Et], the amount of substrate bound by the enzyme at any given time is negligible compared with the total [S]

Question 66

V_0, initial rate, initial velocity, steady state

Answer 66

* In a typical reaction, [S] may be five or six orders of magnitude higher than [E] * If only the beginning of the reaction is monitored (first 60 seconds or less) * changes in [S] can be limited * [S] can be regarded as a constant * V₀ is then the function of [S] * aka the steady state * V₀ = k₂[ES] * k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products

Question 67

V_max, maximum velocity

Answer 67

* At relatively low [S], V₀ increases almost linearly with an increase in [S] * At higher [S], V₀ increases by smaller and smaller amounts in response to increases in [S] * Evenutally increases in V₀ are vanishingly small as [S] increases, because the enzyme is saturated * This plateau-like of the V₀ region is close to the maximum velocity, V_max * V_max occurs when E is saturated (ES = E_t) it is shown as * V_max = k₂[E_t] * k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products

Question 68

K_m, Michaelis constant

Answer 68

* The [S] at which V₀ is half maximal * K_m = 1/2 • V_max * K_m = (K_-1 + K₂) / K₁ * K_m = breakdown / formation * k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products

Question 69

Michaelis-Menten equation

Answer 69

* the rate equation for a one-substrate enzyme-catalyzed reaction * V_0: initial reaction rate * V_max: maximum reaction rate * [S]: initial substrate concentration * K_M: Michaelis constant

Question 70

**E**nzyme-**S**ubstrate complex and **P**roduct formation steps

Answer 70

* k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products * K_-2 = rate constant for the reverse reaction of catalysis

Question 71

enzyme saturation

Answer 71

* At low [S] most of the enzyme is in the uncombined form E * rate is proportional to [S] because * Equilibrium is pushed toward formation of more ES as [S] increases * When virtually all the enzyme is present as the ES complex and [E] is small the enzyme is said to be saturated * maximum initial rate of the catalyzed reaction (V_max) is observed * further increases in [S] have no effect on rate * distinguishing characteristic of enzymatic catalysts * responsible for the plateau in saturation kinetics

Question 72

pre– steady state

Answer 72

* initial period * When the enzyme is first mixed with a large excess of substrate * the concentration of ES builds up * too short to be easily observed, lasting just microseconds

Question 73

steady state

Answer 73

* when [ES] (and the concentrations of any other intermediates) remains approximately constant over time * V₀ generally reflects the steady state, even though V₀ is limited to the early part of the reaction

Question 74

steady-state kinetics

Answer 74

analysis of the initial rates: pre-ready states and steady state

Question 75

Early in the reaction [P] is negligible, and we make the simplifying assumption that the _____ \_\_\_\_\_, can be ignored

Answer 75

* reverse reaction, P → S (k_-2) * k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products * k-₂ = rate constants, EP breakdown to products

Question 76

An important relationship emerges from the Michaelis-Menten equation in the special case when V₀ is exactly one-half V_max

Answer 76

Question 77

all enzymes that exhibit a _____ dependence of V₀ on [S] are said to follow Michaelis-Menten kinetics. The practical rule that ______ when V₀ = ½V_max holds for all enzymes that follow Michaelis-Menten kinetics. The most important exceptions to Michaelis-Menten kinetics are the _____ \_\_\_\_\_

Answer 77

* hyperbolic * K_m = [S] * regulatory enzymes

Question 78

K_m can vary greatly from enzyme to enzyme and even for different ______ of the same enzyme

Answer 78

substrates

Question 79

K_d, dissociation constant

Answer 79

* why is rate-limiting variable removed from the formula? * When k₂ is rate-limiting * k₂ \<\< K_-1 * k₂ * K_m = (K_-1 + K₂) / K1 → K_m = k_-1/k₁ * ​When these conditions hold true, K_m represents a measure of the substrate-binding affinity—does not apply for most enzymes * k₁ = rate constants, ES formation * k_-1 = rate constants, ES breakdown to reactants * k₂ = rate constants, ES breakdown to products

Question 80

When k₂ \>\> K_-1, K_m is

Answer 80

* k₂ \>\> K_-1, * K_m = (K_-1 + K₂) / K1 → K_m = k₂/k₁ * ​K_m DOES NOT represents a measure of the substrate-binding affinity * k1 = rate constants, formation * k-1 = rate constants, breakdown to reactants * k2 = rate constants, breakdown to products

Question 81

When k₂ and K_-1, are comparable, K_m is

Answer 81

* K_m remains a more complex function of all three rate constants * K_m = (K_-1 + K₂) / K1 * ​K_m DOES NOT represents a measure of the substrate-binding affinity * k₁ = rate constants, formation * k_-1 = rate constants, breakdown to reactants * k₂ = rate constants, breakdown to products

Question 82

k_cat, turnover number

Answer 82

* a more general rate constant * describes the limiting rate of any enzyme-catalyzed reaction at saturation * If the reaction has several steps and one is rate limiting, it is equivalent to the rate constant for that limiting step * a first-order rate constant * units: s^-1 * equivalent to the number of substrate molecules converted to product in a given unit of time on a single saturated enzyme

Question 83

experimentally, the K_m for an enzyme tends to be similar to the cellular concentration of its \_\_\_\_\_. An enzyme that acts on a substrate present at a very low concentration in the cell usually has a _____ K_m than an enzyme that acts on a substrate that is more \_\_\_\_\_

Answer 83

* substrate * lower * abundant

Question 84

specificity constant

Answer 84

* best way to compare the catalytic efficiencies of different enzymes or the turnover of different substrates by the same enzyme * compares the ratio k_cat/K_m for the two reactions * rate constant for the conversion of E + S to E + P * second-order rate equation * depends on the concentration of two reactants: [Et] and [S] * units: M^-1s^-1 * ^​has upper limit * 10⁸ - 10⁹ M^-1s^-1 * imposed by the rate at which E and S can diffuse together * many enzymes have a specificy constant near this range * Such enzymes are said to have achieved catalytic perfection

Question 85

Enzymatic reactions with two substrates usually involve transfer of an ____ or a _____ \_\_\_\_\_ from one substrate to the other. These reactions proceed by one of several different pathways. In some cases, both substrates are bound to the enzyme concurrently at some point in the course of the reaction, forming a ____ \_\_\_\_\_\_ complex; the substrates bind in a random sequence or in a specific order. In other cases, the first substrate is converted to _____ and dissociates before the second substrate binds, so no ______ complex is formed

Answer 85

* atom * functional group * noncovalent ternary * product * ternary

Question 86

In the case of bisubstrate reactions, steady-state kinetics can help determine whether a _____ \_\_\_\_\_ is formed during the reaction. In these examples double-reciprocal plots the concentration of substrate 1 is varied while the concentration of substrate 2 is held constant. This is repeated for several values of [S2], generating several separate lines. (a) _____ lines indicate that a ternary complex is formed in the reaction; (b) _____ lines indicate a Ping-Pong (double-displacement) pathway.

Answer 86

* ternary complex * Intersecting * parallel

Question 87

Enzyme inhibitors

Answer 87

* molecules that interfere with catalysis, slowing or halting enzymatic reactions * most important pharmaceutical agents * two broad classes of enzyme inhibitors: reversible and irreversible

Question 88

Three types of reversible inhibition

Answer 88

* Competitive * Uncompetitive * Mixed

Question 89

Reversible Inhibition: competitive inhibitor

Answer 89

* competes w/substrate for the active site of an enzyme * While inhibitor (I) occupies the active site, it prevents binding of the substrate to the enzyme * Many are structurally similar to the substrate and combine with the enzyme to form an EI complex, but without leading to catalysis

Question 90

Reversible Inhibition: competitive inhibitor Competitive inhibition can be analyzed quantitatively by steady-state kinetics. In the presence of a competitive inhibitor, the Michaelis-Menten equation becomes

Answer 90

* αK_m = the K_m observed in the presence of the inhibitor, often called the “apparent” K_m * K_m increases in the presence of inhibitor by the factor α when the [S] is at V₀ = ½V_max * This effect on apparent K_m, combined with the absence of an effect on V_max, is diagnostic of competitive inhibition and is readily revealed in a double-reciprocal plot * KI = equilibrium constant for inhibitor binding,

Question 91

Reversible Inhibition: competitive inhibitor Because the competitive inhibitor binds reversibly to the enzyme, the competition can be biased to favor the substrate simply by

Answer 91

adding more substrate

Question 92

Reversible Inhibition: uncompetitive inhibitor

Answer 92

* observed only with enzymes having two or more substrates * doesn't bind at the substrate active site * binds only to the ES complex * at high [S], V₀ approaches Vmax/α' * lowers the measured V_max * αKm also decreases, because the [S] required to reach one-half V_max decreases by the factor α' * In the presence of an uncompetitive inhibitor, the MichaelisMenten equation is altered to

Question 93

Reversible Inhibition: mixed inhibitor

Answer 93

* observed only with enzymes having two or more substrates * doesn't bind at the substrate active site * binds to either E or ES * usually affects both K_m and V_max * special case of α = α' * rarely encountered in experiments * defined as noncompetitive inhibition * The rate equation describing mixed inhibition is

Question 94

Reversible Inhibition * For all reversible inhibitors, the apparent V_max = \_\_\_\_\_\_, because the right side of the equation always simplifies to _____ at sufficiently high [S] * For competitive inhibitors, α' = _____ and can thus be ignored * αK_m * _​as always, equals the [S] at which V₀ = 1/2 • V_max * or more generally when V₀ = \_\_\_\_\_\_ * This condition is met when [S] = \_\_\_\_\_ * If only one of two reaction products is present, no _____ reaction can take place. However, a product generally binds to some part of the active site, thus serving as an \_\_\_\_\_\_

Answer 94

* V_max/α' * V_max/α' * 1.0 * V_max/2α' * αK_m/α' * reverse * inhibitor

Question 95

Reversible Inhibition: Duble Reciprocal Plots * offers an easy way of determining whether an enzyme inhibitor is competitive, uncompetitive, or mixed * In this example, [E] is constant * 1st set: [S] is held constant to measure the effect of increasing [I] on initial V₀ * 2nd set: [I] is held constant but [S] is varied * Results are plotted as 1/V₀ versus 1/[S]

Answer 95

**Competitive Inhibition** * one set is obtained in the absence of inhibitor * two at different concentrations of a competitive inhibitor * Increasing [I] results in a family of lines with a common intercept on the 1/V₀ axis but with different slopes * Because the intercept on the 1/V₀ axis equals 1/V_max, we know that Vmax is unchanged by the presence of a competitive inhibitor * Regardless of the competitive [I], a sufficiently high [S] will always displace the I from the enzyme’s active site * The value of α can be calculated from the change in slope at any given [I] * Knowing [I] and α, we can calculate K_I in: α = 1 + ([I] / K_I) **Uncompetitive / Mixed Inhibition** * Changes in axis intercepts signal changes in V_max and K_m

Question 96

Irreversible Inhibition: irreversible inhibitors

Answer 96

* two way it functions * bind covalently with or destroy a functional group on an enzyme that is essential for the enzyme’s activity * or form a particularly stable noncovalent association * Noncovalent binding is enough, if that binding is so tight that the inhibitor dissociates only rarely * if one can design a molecule that looks like the reaction transition state, it should bind tightly to the enzyme

Question 97

Irreversible Inhibition Amino acids with key catalytic functions in the active site can sometimes be identified by determining which residue is covalently linked to an ______ after the enzyme is inactivated

Answer 97

inhibitor

Question 98

Irreversible Inhibition: suicide inactivators, mechanism-based inactivators

Answer 98

* relatively unreactive until they bind to the active site of a specific enzyme * undergoes the first few chemical steps of the normal enzymatic reaction, but instead is converted to a very reactive compound that combines irreversibly with the enzyme * hijack the normal enzyme reaction mechanism to inactivate the enzyme

Question 99

transition-state analogs

Answer 99

* stable molecules designed to resemble transition states * bind to an enzyme more tightly than does the substrate in the ES complex, because they fit into the active site better * they form a greater number of weak interactions than the substrate

Question 100

The pH range over which an enzyme undergoes changes in activity can provide a clue to the type of _____ \_\_\_\_\_ _____ involved. In the closely packed environment of a protein, the pKa of amino acid side chains can be significantly \_\_\_\_\_. For example, a nearby positive charge can _____ the pKa of a Lys residue, and a nearby negative charge can _____ it.

Answer 100

* amino acid residue * altered * lower * increase

Question 101

acylation (or alkanoylation)

Answer 101

* the process of adding an acyl group to a compound * compound providing the acyl group is called the acylating agent.

Question 102

regulatory enzyme

Answer 102

* in cellular metabolic activities, many enzymes work together in a sequence to carry out the given metabolic process * in an enzymatic process, were more thatn one steps are involved, the reaction product of one reaction acts as the substrate for the next enzyme * in a multi-step process, there'll be one enzyme responsible for the overall rate of that process, the rate limiting enzyme called the regulatory enzyme * regulary enzymes show enhanced / decreased catalytic activities in response to other molecules (signals)

Question 103

Methods of Enzyme Regulation

Answer 103

* Allosteric Regulation * Reversible Covalent Modifications * Proteolytic Cleavage * Feedback Regulation * Regulation by Isoenzymes

Question 104

Allosteric Regulation / enzymes

Answer 104

* function through reversible, noncovalent binding of **allosteric modulators** or **allosteric effectors** * regulatory compounds * generally small metabolites or cofactors * can be inhibitory or stimulatory * enzymes have additional conformations induced by the binding modulators which can produce more or less active forms of enzyme * enzymes tend to be multisubunit proteins, and in some cases the regulatory site(s) and the active site are on separate subunits * homotrophic enzyme: when the modulator is the substrate itself, the active site and regulatory site are the same * heterotropic enzyme: when the modulator is a molecule other than the substrate * Enzymes with several modulators generally have different specific binding sites for each

Question 105

Allosteric Regulation / enzymes allosteric modulators should not be confused with _____ and _____ inhibitors. Although the latter bind at a second site on the enzyme, they do not necessarily mediate _____ changes between active and inactive forms, and the _____ effects are distinct

Answer 105

* uncompetitive * mixed * conformational * kinetic

Question 106

Reversible Covalent Modification

Answer 106

* enzymes tend to be multisubunit proteins, and in some cases the regulatory site(s) and the active site are on separate subunits * involves the addition or removal of some type of group, most commonly the phosphoryl group, onto or from the enzyme to modify the activity of enzymes and many other proteins * addition of phosphoryl groups involves the use of ATP (energy source) and requires a protein called protein kinase * phosphatases are proteins that can remove phosphoryl groups from enzymes * the addition or removal of phosphoryl groups is reversible.

Question 107

Proteolytic Cleavage

Answer 107

* enzymes are produced as inactive forms called a zymogen or pre-enzyme * active site will be masked/covered by part of the polypeptide chain, "inactivating" the zymogen * zymogens are converted to active enzymes by the removal of specific parts of enzyme by proteolytic cleavage * specific cleavage causes conformational changes that expose the active site of enxyme * irreversible

Question 108

Feedback Regulation

Answer 108

* not feedback inhibition * the end product of an enzymatic pathway directly inhibit the synthesis of another enzyme by interfering with the gene of that enzyme * enzyme is not directly inhibited by the end product * end product reduces the concentration of enzyme by inhibiting the synthesis of new enzyme molecules

Question 109

Regulation by Isoenzymes

Answer 109

* not a direct method of enzyme regulation * enzymes doing similar catalytic function but have different amino acid sequences and kinetic parameters, the K_m, V_max and V₀ differ

Question 110

Salt bridges

Answer 110

* in proteins are bonds between oppositely charged residues that are sufficiently close to each other * a non-covalent interaction between two ionized sites * has two components: a hydrogen bond and an electrostatic interaction * a proton migrates from a carboxylic acid group to a primary amine or to the guanidine group in Arg. T * ypical salt bridges involve Lys or Arg as the bases and Asp or Glu as the acids. * Of all the non-covalent interactions, salt bridges are among the strongest.

Question 111

Allosteric enzymes show relationships between V₀ and [S] that differ from Michaelis-Menten kinetics. The plots of V₀ versus usually produce a ______ \_\_\_\_\_ curve, rather than the ______ curve typical of nonregulatory enzyme. The symbol _____ or _____ is often used to represent the substrate concentration giving half-maximal velocity of the reaction catalyzed by an allosteric enzyme

Answer 111

* sigmoid saturation * hyperbolic * [S]_0.5 * K_0.5

Question 112

Sigmoid kinetic behavior generally reflects cooperative interactions between multiple protein \_\_\_\_\_\_. Changes in the structure of one subunit are translated into structural changes in adjacent subunits, an effect mediated by _____ \_\_\_\_\_ at the interface between subunits

Answer 112

* subunits * noncovalent interactions

Question 113

Answer 113

* The sigmoid curve of a homotropic enzyme, in which the substrate also serves as a positive (stimulatory) modulator, or activator * resemblance to the oxygen-saturation curve of hemoglobin * The sigmoidal curve is a hybrid curve in which the enzyme is present primarily in the relatively inactive T state at low substrate concentration, and primarily in the more active R state at high substrate concentration. * The curves for the pure T and R states are plotted separately in color

Question 114

Answer 114

* The effects of several different concentrations of a positive modulator (+) or a negative modulator (-) on an allosteric enzyme in which K0.5 is altered without a change in Vmax. * The central curve shows the substrate-activity relationship without a modulator.

Question 115

Answer 115

A less common type of modulation, in which V_max is altered and K_0.5 is nearly constant

Question 116

In another important class of regulatory enzymes, activity is modulated by _____ \_\_\_\_\_\_ of one or more of the amino acid residues in the enzyme molecule. Common modifying groups include phosphoryl, acetyl, adenylyl, uridylyl, methyl, amide, carboxyl, myristoyl, palmitoyl, prenyl, hydroxyl, sulfate, and adenosine diphosphate ribosyl groups are generally linked to and removed from a regulated enzyme by _____ \_\_\_\_\_\_.

Answer 116

* covalent modification * separate enzymes

Question 117

Introduction of a charge can alter the local properties of the enzyme and induce a change in \_\_\_\_\_\_. Introduction of a hydrophobic group can trigger association with a \_\_\_\_\_\_

Answer 117

* conformation * membrane

Question 118

\_\_\_\_\_ is the most important type of regulatory modification. Phosphoryl Groups Affect the _____ and _____ \_\_\_\_\_ of Enzymes. it can have dramatic effects on enzyme _____ and thus on _____ \_\_\_\_\_

Answer 118

* Phosphorylation * Structure * Catalytic Activity * conformation * substrate binding

Question 120

The attachment of phosphoryl groups to specific amino acid residues of a protein is catalyzed by _____ \_\_\_\_\_. In the reactions, the y-phosphoryl group derived from a nucleoside triphosphate (usually ATP) is transferred to a particular Ser, Thr, or Tyr residue (occasionally His as well) on the target protein. This introduces a bulky, charged group into a region of the target protein that was only moderately polar. When the modified side chain is located in a region of an enzyme critical to its three dimensional structure, phosphorylation can have dramatic effects on enzyme _____ and thus on _____ \_\_\_\_\_ and \_\_\_\_\_\_

Answer 120

* protein kinases * conformation * substrate binding * catalysis

Question 121

Removal of phosphoryl groups from these same target proteins is catalyzed by _____ \_\_\_\_\_

Answer 121

protein phosphatases

Question 122

Phosphorylation of an enzyme can affect catalysis in another way: by altering \_\_\_\_\_\_-\_\_\_\_\_\_ \_\_\_\_\_

Answer 122

substrate-binding affinity

Question 123

The Ser, Thr, or Tyr residues that are typically _____ in regulated proteins occur within common structural motifs, called consensus sequences, that are recognized by specific _____ \_\_\_\_\_\_.

Answer 123

* phosphorylated * protein kinases

Question 124

Amino acid sequence is not the only important factor in determining whether a given residue will be \_\_\_\_\_\_, however. Protein folding brings together residues that are distant in the primary sequence; the resulting three-dimensional structure can determine whether a protein kinase has access to a given residue and can recognize it as a \_\_\_\_\_\_. Another factor influencing the substrate specificity of certain protein kinases is the proximity of other _____ \_\_\_\_\_\_

Answer 124

* phosphorylated * substrate * phosphorylated residues

Question 125

Regulation by phosphorylation is often complicated. Some proteins have consensus sequences recognized by several different protein kinases, each of which can phosphorylate the protein and alter its enzymatic activity. In some cases, phosphorylation is \_\_\_\_\_\_: a certain residue can be phosphorylated only if

Answer 125

* hierarchical * a neighboring residue has already been phosphorylated

Question 126

For some enzymes, an inactive precursor called a ______ is cleaved to form the active enzyme. Many proteolytic enzymes (\_\_\_\_\_) of the stomach and pancreas are regulated in this way. Chymotrypsin and trypsin are initially synthesized as chymotrypsinogen and trypsinogen. Specific cleavage causes _____ \_\_\_\_\_\_ that expose the enzyme active site. Because this type of activation is \_\_\_\_\_, other mechanisms are needed to inactivate these enzymes. Proteases are inactivated by _____ \_\_\_\_\_ that bind very tightly to the enzyme active site

Answer 126

* zymogen * proteases * conformational changes * irreversible * inhibitor proteins

Question 127

Proteases are not the only proteins activated by \_\_\_\_\_. In other cases, however, the precursors are called not zymogens but, more generally, _____ or \_\_\_\_\_, as appropriate. For example, the connective tissue protein collagen is initially synthesized as the soluble precursor procollagen

Answer 127

* proteolysis * proproteins * proenzymes