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Flashcards in Enzymes Deck (155):
1

Highly specific, extremely fast, biological catalysts

-Proteins

Enzymes

2

Mutations in proteins and enzymes are the cause of many

Diseases

3

Enzymes can be used as drugs, an example of this is the clot buster

Activase

4

Enzymatic reactions are multi-step reactions. The first step in an enzyme reaction is the enzyme binds

Substrate (forms the ES complex)

5

The second step in an enzyme reaction is the

conversion of ES to EP

6

The third step of an enzymatic reaction is the

Dissociation of P and regeneration of E

7

A molecule that accelerates a chemical reaction and is regenerated at the end of the reaction

Catalyst

8

Says that an enzyme has an active site that fits only a specific substrate

-ex: yeast fermented the D- but not L- forms of glucose, mannose, and galactose

Fisher's lock and key model

9

A three dimensional cleft formed by catalytic amino acids that come from different parts of the protein sequence

Enzymes active site

10

Bind substrates but don't carry out a chemical reaction

Receptors

11

Bind substrates and carry out chemical reactions

Enzymes

12

Close together in tertiary structure, but not in primary structure

Catalytic amino acids

13

Only a very small portion of an enzymes aminos function in the active site, the rest serve as a

Scaffold

14

Small pockets lined with a few catalytic amino acids that participate in substrate binding and catalysis

Active site

15

Small part of the total volume of the enzyme and is non-polar (excludes water unless water is a reactant)

Active site

16

The nonpolar characteristics of active sites enhances substrate binding by increasing

Electrostatic interactions

17

Substrates bind active sites by way of many

weak non-covalent interactions

18

What is the energy for a

1.) Covalent interaction?

2.) Non-covalent interaction

1.) 50 kcal/mol

2.) 0.5-2 kcal/mol

19

Enzymes can undergo many catalytic cycles because substrates and products bind

reversibly

20

Precise active site conformation of glucokinase explains specific binding and reaction of ATP with glucose but not galactose. Why does galactose not bind even though the two differ by only a single hydroxyl?

Cooperative binding

21

The lock and key model proved to be inadequate when looking at glucokinase and hexokinase because

Since they bound glucose, they should have been able to bind water and react with ATP, but they could not

22

The lock and key model failed to explain why ATP did not react with water, which paved the way for the

Induced-fit model

23

Summarize the induced fit model

Substrate binding causes a conformational shift in the enzyme, which stabilizes the active conformation and allows catalysis

24

In the induced fit model, a specific substrate activates the enzyme by

Orienting catalytic groups on the enzyme

25

What was the experimental evidence supporting the induced fit model?

The binding of glucose induced a large conformational change in glucokinase

26

In the case of ATP transferring a phosphate to NMP rather than water, we see that the induced fit conformational change assures that a catalytically competent complex is formed only when

-prevents the reaction with water

Both ATP and NMP are bound to NMP kinase

27

Enzymes increase the rates of reactions by a factor of

10^6-10^17 times

28

The minimum energy required for two molecules to react

-inversely proportional to reaction rate

Activation energy

29

Enzymes work by decreasing the

Activation barrier (transition state energy)

30

Enzymes have no effect on the

Gibbs free energy

31

Enzymes create a new reaction pathway with a lower activation energy through specific binding to the

Transition state

32

How do enzymes lower the transition state energy?

Tighter binding to transition state than to substrate

33

What are five ways enzymes speed up the rate of a reaction?

1.) Proximity (all)
2.) Stabilization of transition state (all)
3.) Covalent or nucleophilic catalysis (some)
4.) Acid-base catalysis (most)
5.) metal ion catalysis (many)

34

Increase the effective concentration of reactants that are in the proper orientation

Proximity

35

In a chain of successive reactions, the rate enhancement due to proximity increases as the

Products structure becomes more rigid (less ability to be in improper orientation)

36

What are the three enzymatic strategies employed by the protease enzyme chymotrypsin?

1.) Nucleophilic or covalent catalysis
2.) General acid-base catalysis
3.) Transition state stabilization (oxyanion hole)

37

The serine residue in the active sight of chymotrypsin is a powerful

Nucleophile (but must be deprotonated)

38

The pKa of the side chain of serine is

pKa = 13

39

The serine residue in the active site of chymotrypsin is turned into a potent nucleophile by deprotination using

Acid-base catalysis

40

The extremely reactive serine residue in chymotrypsin is created by the

Catalytic triad (Ser, His, Asp)

-His accepts a proton to become HisH+

41

In the chymotrypsin active site, once serine is deprotonated, it

Nucleophilically attacks the carbonyl carbon of amino acid, forming the tetrahedral intermediate (oxyanion hole), which then reforms the double bond and kicks off the amine

42

Serine nucleophilically attacking the carbonyl carbon is an example of

Covalent Catalysis

43

Binds the transition state tightly and stabilizes the transition state by way of H-bonds which were not possible in the reactant

Oxyanion hole

44

A space in the enzyme active site that is ready to bind a negatively charged group

Oxyanion hole

45

Summarize how chymotrypsin cleaves at the C-terminal end of the aromatic amino acids (Phe, Tyr. Trp)

Aspartate H-bonds w/ histidine, the His Nitrogen attacks the serine OH,allowing the SerO- to nucleophilically attack the carbonyl carbon of the amino, forming the tetrahedral intermediate. The oxyanion hole stabilizes the intermediate until the carbonyl is reformed, breaking the amide bond and the amine fragment is released. His then takes a proton from water, which generates OH, which attacks the carbonyl carbon of the residue still bound to serine, forming the tetrahedral intermidiate. The electrons reform the double bond, releasing a now carboxylic acid from serine. Serine takes a proton back from histidine.

46

A reaction where ΔG

Exergonic Reaction

47

A reaction where ΔG > 0

-Non spontaneous reaction

Endergonic reaction

48

An endergonic reaction can be driven forward by

Coupling it to an exergonic reaction

49

What is the ΔG˚ of ATP hydrolysis?

-30.5 kJ/mol

50

What are three reasons that ATP hydrolysis is so favorable?

1.) ADP and Pi (products) are more stable than ATP (reactant)
2.) Electrostatic repulsion (ADP has -2 charge and ATP has -3)
3.) Resonance stabilization of Pi

51

What is a great example of a coupling reaction?

Coupling the phosphorylation of Glucose to form glucose-6-phosphate with ATP hydrolysis

52

The ΔG˚ value provides absolutely no information about

Reaction rates

53

Kcat is the rate constant of

ES ---> E + P

54

The velocity of an enzyme reaction is proportional to the

Concentration of the ES complex i.e. [ES]

55

How can we use the MM equation to determine Kcat and Km?

Measure velocity of the reaction

56

How do we measure the velocity (speed) of the reaction?

Measure the change in concentration of substrate or product over the time of the enzymatic reaction

57

The MM equation shows that enzymatic rate/ velocity increases linearly with

Enzyme concentration

58

The MM equation shows that enzymatic rate increases asymptomatically with

Increasing [S]

59

On a plot of velocity vs [S], Km can be thought of as

Vmax/2

60

On a plot of velocity vs [S], Kcat can be thought of as

Vmax/[E]

61

The ratio of Kcat/Km is called the

-measure of enzyme efficiency

Specificity constant

62

From a definition standpoint, we can think of Km as being inversely proportional to the

The enzymes affinity for substrate

-Large Km = small affinity

-Small Km = Large affinity

63

Result in a high specificity constant

1.) Rapid Turnover (large kcat)
2.) High enzyme affinity for substrate (Low Km)

64

A measure of the affinity of the substrate for the enzyme

Km

65

Characteristic for an enzyme for a particular substrate

-Changes with pH or Temperature

Km

66

Km values are usually close to

Physiological substrate concentration

67

The maximum velocity with which the enzyme can catalyze the reaction

Vmax (units = M/s)

68

The turnover number of an enzyme, representing the number of moles of substrate converted to products persecond per mol of enzyme

Kcat (units s^-1)

69

Vmax is linearly dependent on

[E]

70

Some enzymes require cofactors. An enzyme and it's cofactor are called a

Holoenzyme

71

Small organic molecules, mostly derived from vitamins, that are either noncovalently bound to the enzyme, or covalently bound (prosthetic groups)

Coenzymes

72

Apoenzymes without their cofactor are

Inactive

73

Provide the functionalities not found in the natural amino acids

Coenzymes

74

Have different primary structures or amino acid sequence, but catalyze the same chemical reaction and act upon the same substrate

-Thought to have evolved from gene duplication and divergence

Isozymes

75

Allow fine tuning of metabolism to meet the needs of a given tissue or developing stage

Isozymes

76

Do isozymes have the same Km and kcat as their enzyme isomers?

No

77

Measurement of isozyme levels helps in

diagnosis

78

What happens within one day of heart muscle damage?

-indicates myocardial infarction

1.) LDH isozyme H4 increases within 12-24 hrs.
2.) Level of MB creatine kinase in the blood increases

79

Enzyme activity is affected by temperature. Explain how temperature increases rate.

As temp increases, there are more molecular collisions between E and S

80

Enzyme activity is affected by temperature. Explain how temperature decreases rate.

When temp gets too high, proteins are denatured, causing the decrease in rate

81

What are the two negative effects that pH can have on an enzyme?

1.) extreme pH's irreversibly denature the enzyme and activity is lost forever
2.) Can reversibly affect the enzyme by ionizing the functional groups involved in catalysis

82

Enzymes in different organs have different pH optimum, but most intracellular enzymes have a pH optimum of

pH = 7.2

83

Enzyme concentration is regulated by

1.) Synthesis (transcription, translation, etc.)
2.) degradation (proteolysis)

84

Enzyme activity is regulated by compartmentalization in

Cell organelles

85

Enzyme activity is regulated by post translational modifications such as

Phosphorylation, glycosylation, methylation, etc.

86

Enzyme activity is regulated by regulatory proteins such as

Transcription Factors

87

Enzyme activity is inhibited by

Feedback inhibitors, enzyme inhibitors, and products

88

ADP-ribosylation of EF2 blocks its capacity to carry out translocation of the growing polypeptide chain, thus

-Accounts for remarkable toxicity of diptheria toxin

Protein synthesis ceases

89

Over 95% of protein phosphorylation occurs on

Serine resiues

90

Highly negatively charged phosphoryl group can alter

Substrate binding and catalytic activity

91

Is reversible and fast and it can either activate or inhibit the enzyme

Phosphorylation

92

Abnormal levels of protein phosphorylation are a cause or consequence of major diseases such as

Cancer, diabetis, and arthritis

93

What is an irreversible way to activate enzymes?

Proteolysis

-apoptosis mediated by caspases from procaspases

94

A prime example of feedback regulation of an enzyme is that of the allosteric enzyme Aspartate Transcarbamylase, which is activated by? inhibited by?

Activated by ATP

Inhibited by CTP

95

Enzymes that change their conformational ensemble upon binding of an effector, which results in an apparent change in binding affinity at a different ligand binding site.

Allosteric Enzymes

96

All allosteric enzymes are

Oligomeric

97

Allosteric molecules (bind in the allosteric site) do not resemble the

Substrate

98

Allosteric enzymes can be distinguished kinetically because they show

Sigmoidal curves

99

Do allosteric enzymes follow MM kinetics?

No

100

Enzymes following MM kinetics show a

Hyperbolic curve

101

Allosteric curves are sigmoidal due to the allosteric enzyme's

Cooperative binding

102

Allosteric enzymes exist in which two equilibrium conformations?

1.) Active (R) state
2.) Inactive (T) state

103

Binds substrate better and/or has higher catalytic activity

R-state

104

How does an allosteric activator increase substrate binding and/or activity?

Binds to the allosteric site and stabilizes the R-state

105

How does an allosteric inhibitor decrease substrate binding and/or activity?

Binds to the allosteric site and stabilizes the inactive T-state

106

In AcT, the inactive T-state is favored by

CTP binding

107

Inhibitors shift the allosteric sigmoidal curve to the?

Right

108

Activators shift the allosteric sigmoidal curve to the?

Left

109

The basis for clinical drug therapies: antiviral, antibacterial, and antitumor therapies

Enzyme inhibition by small molecules

110

What are the two main classes of inhibitors?

1.) Irreversible inhibitors (bind covalently)
2.) Reversible inhibitor (bind reversibly)

111

What are the three types of reversible inhibitors?

1.) Competitive
2.) Noncompetitive
3.) Uncompetitive

112

Substrate analogs that form a covalent bond with and active site amino acid

-ex: penicillin bonding to transpeptidase

Irreversible inhibitors

113

Aspirin is an example of an

Irreversible inhibitor

114

Bind the same active site as the substrate

Competitive inhibitor

115

Bind to a site separate from the active site

Noncompetitive inhibitor

116

Stable molecules that resemble geometric and/or electronic features of the highly unstable transition state.

-Potent inhibitors

Transition state analogues

117

What are the kinetic features of a competitive inhibitor?

-Vmax is unchanged
-Km increases

No inhibition at high [S]

118

What are the kinetic features of a noncompetitive inhibitor?

-Km is unchanged
-Vmax decreases

-High [S] does not overcome inhibition

119

Where do the lines intersect on a linweaver-burke plot for

1.) competitive inhibition?
2.) noncompetitive inhibition?

1.) y-axis
2.) negative x-axis

120

The most widely prescribed drug that lowers cholesterol

Statins (competitive inhibition)

121

Statins act by inhibiting

HMG-CoA reductase

122

What is Pi?

Inorganic Phosphate

123

The gibbs free energy represents the

Maximal work under a set of conditions

124

ATP is so reactive because it has three phosphates and a charge of

-3 or -4, averaging to -3.5

125

Where do coupled reactions occur in the enzyme?

On the same active site

126

The dissociation constant of [ES]

Kcat

127

What are the trends of the [ES] dissociation constant Km?

The smaller the Km, the faster the reaction typically is

-Small Km means enzyme does not dissociate from [ES] and reaction proceeds

128

What is the rate limiting step of an enzymatic reaction?

Kcat

129

When [S] is in excess, the velocity increases linearly with

[E]

130

Why does the rate increase asymptomatically with increasing substrate concentration?

At some point all of the enzyme active sites are saturated with substrate, and Vmax has been reached

131

The higher the value of Km, the

slower the reaction

132

Mutations in enzymes that affect the ability of substrate to bind will also effect

Km

133

What are the units of Km?

M, mM, or uM

134

Is roughly equal to the physiological substrate concentration

Km

135

What is the fastest known enzyme?

-Called the perfect enzyme

Carbonic anhydrase

136

An inactive enzyme without its cofactor

Apoenzyme

137

Allow variability that expands beyond that of the 20 amino acids in enzymes

Coenzymes

138

What is a common example of isozymes?

Hexokinase and Glucokinase

139

The liver synthesizes and utilizes

Glucokinase

140

The optimum temperature of human enzymes is

38 degrees celcius

141

Km and Kcat values are affected by

Temperature and pH

142

Why is pH so important to enzymes?

The H-bonds that form from the catalytic aminos will not be possible if pH is outside of optimum range

143

We don't want to have to synthesize an enzyme every time we need it, because that will take to long. So, we synthesize enzymes and keep them in the inactive form called a

Zymogen

144

The activation of a zymogen to an enzyme is

Irreversible

145

Can be used to turn an enzyme off by inactivating a Ser, Thr, or Tyr in the active site

Phosphorylation

146

An example of enzyme regulation by specific proteolysis

Zymogen activation

147

Regulation of an enzyme at a distance (how many metabolic pathways are regulated)

Allosteric Regulation

148

All allosteric enzymes are oligomeric, meaning they are

Dimers, trimers, tetramers, etc

149

Do not look like the enzymes substrate, thus they do not bind to the active site. Instead, they bind to a regulatory site and cause some form of conformation change in the enzyme which either activates or inhibits the enzyme

Allosteric Effectors

150

Their association with an enzyme is not allosteric because they actually bind to the active site

Cofactors

151

Instead of a Km, allosteric enzymes have a

K0.5

152

What type of allosteric inhibitor or activator affects K0.5 but not Vmax?

K-type

153

What type of allosteric inhibitor or activator affects Vmax but not K0.5?

V-type

154

AcT is a hexamer, the active site is at the interface of two subunits in the T-state. When an activator is added, the interface is broken, freeing the active sites and stabilizing the

R states

155

Most drugs are

Inhibitors that target enzymes

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