Exam 2 (Lectures 11-13)

Question 1

First Order Kinetics

Answer 1

there is only one reactant that is being worked on by the enzyme (unimolecular reaction)

v = k [A]

[=] s^-1

Question 2

Second Order Kinetics

Answer 2

deals with 2 substrates to combine into 1 product

v = k [A][B}

[=] mol^-1s^-1

Question 3

Pseudo-First Order Kinetics

Answer 3

bimolecular reaction with one reactant far in excess

rate of reaction depends on the concentration of only one substrate (limiting reactant)

Question 4

Zero Order Kinetics

Answer 4

steady state

the reaction is achieved at equilibrium

Question 5

Carbonic Anhydrase

Answer 5

catalyzes hydration of CO₂ to carbonic acid.

proceeds at moderate rate without the enzyme

can increase turnover rate to 10⁶ mol/s

Question 6

Zn⁺² Metal Cofactor

Answer 6

4 coordination sites: 3 Histidine groups and water

destabilizes water and allows loss of proton and quickly converts to OH^-

Question 7

Protons diffuse too slowly to accomodate the turnover rate speed of 10^-6s^-1 so in order for catalysis to continue:

Answer 7

shuffling protons to a buffer of pH 7

Question 8

Enzyme must be regulated to:

Answer 8

allow for optimal functioning to accomodate different metabollic needs

Question 9

Ways to regulate Enzymes

Answer 9

Allosteric Control

Multiple Isozymes

Reversible Covalent Modification

Proteolytic Activation

Controlling Enzyme Levels

Question 10

Allosteric Control

Answer 10

involves a regulatory sites other than the active site to which a small molecule can bind.

coopererativity beetween subunits within the same protein

Question 11

Binding at a small molecule often results in a…

Answer 11

3-dimentional change in the conformation of the enzyme.

Question 12

Cyclic AMP (cAMP)

Answer 12

a small molecule that is formed by cyclization of ATP

an important intracellular messenger and allosteric factor

Question 13

Binding of regulatory small molecules

Answer 13

1. Binding of cAMP to R (regulatory region)
2. Confomational change in R
3. Release of inhibiting R pseudosubstrate sequences
4. 2 freed (active) protein kinase A molecules

Question 14

Catalytic regions (Protein Kinase A)

Answer 14

phosphorylates target protein (enzymes) to either Activate or De-activate it

Question 15

Reciprocal Regulation

Answer 15

two pathways (synthesis and degredation) of the same molecule are not working at the same time to the same extent

Question 16

Protein Phosphatase

Answer 16

the enzyme that dephosphorylates a protein by hydrolyzing the phosphate from the target protein

Question 17

Isozyme

Answer 17

different form of an enzyme, in the same individual, that catalyzes the same reaction but has slightly different amino acid sequence and usually different properties

Question 18

Isozymes may be different in..

Answer 18

the mode of regulation

expression patterns

kinetic parameters

Question 19

Lactate Dehydrogenase Enzyme

Answer 19

tetramer that is made up of H subunit and M subunit

Question 20

H Subunit

Answer 20

higher afinity for the substrate that does M subunit

prevalent in heart because it can function very well in aerobic situation where plenty of oxygen is available

Question 21

M Subunit

Answer 21

is negatively regulated by its product while H subunit is not

prevalent in muscle and liver because it can function very well in anaerobic situation

Question 22

Native Gel Electrophrysis

Answer 22

Proteins are in their natural 3D conformation to preserve tertiary and quaternary structures

Question 23

Thickness of the bands in gel electrophoresis

Answer 23

indicative of how much of a particular subunit is found in that particular sample

Question 24

Reversible Covalent Modification

Answer 24

happens after the protein is translated

phosphorylation, acetylation, myristoylation, ADP ribosylation, Farnesylation, y-carboxylation, suflation, ubiquitination

Question 25

Proteolytic Activation

Answer 25

cleavage of inactive zymogen to activate enzymes (happens to proteosys)

Question 26

Zymogens

Answer 26

Pepsinogen (from stomach), Chymotrypsinogen (pancreas), Trypsinogen (pancreas), Procarboxypeptidase (pancreas), Proelastase (pancreas). they are then transported to the duodenum where they are activated through proteolytic cleavage

Question 27

Proteolytic Activation: Digestive enzymes

Answer 27

proteolytic cascade which begins with cleaving of trypsinogen by enteropeptidase, resulting in active trypsin which activates a digestive protease cascade

Question 28

Proteolytic Activation: Clotting factors

Answer 28

proteolytic cascade resulting in cleaving fibrinogen to form fibrin to form blood clots

Question 29

Proteolytic Activation: Peptide hormones

Answer 29

pro-insulin to insulin

Question 30

Proteolytic Activation: Collagen

Answer 30

procollagen (solluble precursor) to extracellular matrix protein, collagen

Question 31

Proteolytic Activation: Tissue remodeling enzymes

Answer 31

active enzymes that digest tissue, procollagenase to collagenase

Question 32

Proteolytic Activation: Apoptotic enzymes

Answer 32

formation of proteases called caspases from procaspases. caspase activation results in a tightly regulated cell death called programmed cell death or apoptosis.

Question 33

Matrix

Answer 33

innermost portion of the mitochondrion and is the site of many biosynthetic and catabolic reactions

Question 34

Heme Synthesis

Answer 34

formation of protoporphyrin IX (asymmentric ring structure) and chelation of iron

Question 35

Heme synthesis requires...

Answer 35

4 Mitochondrial and 4 Cytosolic enzymes

Question 36

Heme Synthesis: Reaction 1

Answer 36

condensation between Succinyl CoA and Glycine to form d-Aminolevulinate (d-ALA) also a rate limiting step (takes the longest time and dictates how fast or slow the whole pathway will go) Enzyme: d-Aminolevulinate Synthase Site: Mitochondrial Matrix

Question 37

Heme Synthesis: Regulation Reaction 1

Answer 37

Primarily by Negative feedback through heme

Question 38

Heme Negative feedback effect on Regulation

Answer 38

inhibiting transport of d-ALA synthase into the mitochondrion by high heme concentration

Question 39

Where is d-ALA synthase synthesized?

Answer 39

in the cytosol, however it is used in the mitochondrion

Question 40

Heme negative feedback effect on:

Answer 40

transcription of d-ALA synthase, mRNA stability, and translation

Question 41

Heme Synthesis: Reaction 2

Answer 41

Dehydration synthesis where d-ALA is transprted to cytosol and 2 d-ALA condeses to form each porphobilinogen molecule

Question 42

Heme Synthesis: Reaction 3

Answer 42

condensation reaction in which 4 molecules of porphobilinogen condense head to tail to form a linear tetrapyrrole molecule with release of one ammonium ion for each methylene bridge made Enzyme: Porphobilinogen Deaminase

Question 43

Heme Synthesis: Reaction 4

Answer 43

Cyclization: synthesis of an asymmetric ring Uroporphyrinogen III (constructs one of the rings to have the substituents flipped compared to the rest) Enzyme: Uroporphyrinogen III Synthase

Question 44

Ferrochelatase

Answer 44

the enzyme that chelates iron to form heme

Question 45

Heme Synthesis: Reaction 5

Answer 45

forms methyl and vinyl side chains and alters saturation of ring constituents ending with protoporphyrin IX. Site: Mitochondrion

Question 46

Heme Synthesis: Final Reaction

Answer 46

Chelation of iron and positioned into the ring by the enzyme ferrochelatase

Question 47

Chelation

Answer 47

multiple bonds are made between a group and a central metal (in heme: between the pyrrol nitrogens and central iron)

Question 48

Porphyrias

Answer 48

problems with Heme Synthetic pathway 1. Acute intermittent porphyria 2. Congenital erythropoietic porphyria

Question 49

Acute Intermittent Porphyria

Answer 49

deficiency in the enzyme that catalysis reaction 3: Porphobilinogen Deaminase Deficiency Excess porphobilinogen and d-ALA

Question 50

Symptoms of Acute Intermittent Porphyria

Answer 50

severe abdominal pain and neurologic involvement

Question 51

Congenital Erythropoietic Porphyria

Answer 51

Reaction 4: Uroporphyrinogen III Synthase Deficiency. Causes uroporphyrinogen I to be formed in addition to III

Question 52

Symptoms of Congenital Erythropoietic Porphyria

Answer 52

photosensitivity and red fluorescent teeth

Question 53

What is the RBCs average life span?

Answer 53

120 days

Question 54

Primary site of RBC scavenging (filtering out of the bloodstream)

Answer 54

Spleen

Question 55

Hemoglobin Catabolism

Answer 55

1. hemoglobin protein components, globin chains, are hydrolyzed to individual amino acids 2. the iron is recycled 3. heme is broken down in several steps to form 1-stercobilin (excreted in poop) and 1-urobilin (excreted in urine)

Question 56

Heme Oxygenase

Answer 56

breaking of alpha-methene bridge resulting in the release of Fe³⁺ Products are Biliverdin, CO, H₂O, NADP⁺

Question 57

Biliverding Reductase

Answer 57

reduction of central methene bridge to form Bilirubin and NADP⁺

Question 58

Bilirubin is transported from...

Answer 58

the spleen to the bloodstream then the liver but it is poorly soluble (hydrophobic)

Question 59

Bilirubin is not very hydrophilic, what carrier is needed to transport it from the spleen to the bloodstream

Answer 59

Serum Albumin

Question 60

Glucuronyl transferase (liver enzyme)

Answer 60

the enzyme that catalyzes the transfer of glucuronide to bilirubin

Question 61

Bilirubin Diglucuronide

Answer 61

is made by transferring 2 activated glucuronide sugars to 2 propionate side chains of bilirubin it is now hydrophilic enough to be transported in bile through bile duct to the small intestine for further processing

Question 62

Conjugated bilirubin

Answer 62

when glucuronyl transferase does its thing in the liver to form bilirubin diglucuronide synthesized in and released by the liver

Question 63

once the conjugated bilirubin is in the intestine..

Answer 63

the glucuronides are removed and further breakdown carried out by gut bacteria Product: urobilininogen

Question 64

Urobilininogen (orange yellow color)

Answer 64

some is reabsorbed and transported into the kidneys and then excreted in the urine. the remaining is further broken down into 1-stercobilin and excreted in feces

Question 65

Unconjugated bilirubin

Answer 65

bilirubin that came straight from the spleen formed in the spleen

Question 66

Tests for Serum Bilirubin Levels

Answer 66

1. Indirect (unconjugated) bilirubin 2. Direct (conjugated bilirubin)

Question 67

1. Indirect Test for bilirubin

Answer 67

increased unconj. bilirubin results in: Anemia neonatal jaundice resolution of large hematoma

Question 68

Increased of the levels of Unconj. bilirubin is caused by...

Answer 68

the body trying to get rid of a lot of heme all at once overwhelming the system, generally caused by an increased in red blood cell lysis

Question 69

Anemia

Answer 69

can result in the rupturing of RBCs and a lot of heme must be prossesed

Question 70

Neonatal Jaundice

Answer 70

a lot of bilirubin must also be processed at once and can build up in the tissues UV light can make bilirubin go in a conformational change making it easier to extract out of the body

Question 71

Why do bruises change color from red to green to yellow?

Answer 71

because it is the process of going from heme to biliverdin to biliruben

Question 72

2. Direct Test for bilirubin

Answer 72

increased conj. bilirubin liver disease: bile duct obstruction cirrhosis: scarring of the liver hepatitis: infection of the liver

Question 73

Increased of the levels of Conj. biliruben is caused by...

Answer 73

the liver itself due to the liver being unable to dispose of it normally through the bile

Question 74

Total Serum Biliruben

Answer 74

measures both indirect and direct serum bilirubin

Question 75

Enzymes

Answer 75

are catalysts that specifically recognize their binding partners through complementarity of 3-D structure

Question 76

Most enzymes are:

Answer 76

proteins and there are some that are RNA that functions as a catalyst

Question 77

What do enzymes do to the transition state?

Answer 77

they stabilize the transition state by decreasing the activation energy required to form it

Question 78

ΔG^‡

Answer 78

is the activation energy, the amount of energy needed to initiate the reaction

Question 79

Enzyme function:

Answer 79

1. Bring reactants and cofactors close enough and in the correct orientation long enough for reaction to occur 2. Provide an optimal environment for catalysis: a. Supply residues that participate in the reaction: i. act as proton donor/acceptor in acid/base catalysis ii. acidify functional groups in a reactant iii. stabilize a developing charge in a reactant b. Provide torsional energy that allows catalysis: i. conformational change upon reactant or cofactor binding

Question 80

Cofactors can be divided into 2 groups:

Answer 80

Metals - tightly bound, inorganic Coenzymes - small, organic molecules

Question 81

Prosthetic Cofactor Groups

Answer 81

tightly bound cofactors

Question 82

Apoenzyme

Answer 82

an enzyme without its required cofactor

Question 83

Holoenzyme

Answer 83

enzyme with its required cofactor

Question 84

Active Sites

Answer 84

the place a place on the enzyme where the substrate binds

Question 85

Characteristics of the Active Site

Answer 85

1. 3-D cleft or crevice that involves residues from different parts of the protein primary structure 2. Takes up only a small portion of the total enzyme 3. Provides a favorable microenvironment for the reaction 4. Binds substrates through many weak interactions 5. Binding specificity depends on cleft architecture and the presence of amino acids in specific positions

Question 86

Michaelis-Menten Kinetics

Answer 86

enzymes that have a hyperbolic regression that describes its reaction velocity as a function of substrate concentration

Question 87

V_max/2

Answer 87

the velocity at which half of the enzyme binding sites said to be filled with substrate

Question 88

K_M (Michaelis-Menten Constant)

Answer 88

the concentration of substrate at which half of all the possible enzyme binding sites are full [=] micromolar/millimolar

Question 89

The lower the K_M the more...

Answer 89

efficient the binding of the enzyme is for that substrate

Question 90

the higher the V_max, the more...

Answer 90

efficient the enzyme and substrate are at producing the product

Question 91

V_o =

Answer 91

( [S] V_max ) / ( [S] + K_M ) [S] - substrate concentration

Question 92

Lineweaver-Burk Plot

Answer 92

a double reciprocal plot of the original regression function where the y and x axes are 1/V_o and 1/[S] respectively.

Question 93

In a Lineweaver-Burk Plot, where in the plot are K_M and V_max

Answer 93

the x-intercept is equal to -1/K_M while the y-intercept is equal to 1/V_max

Question 94

Reversible Inhibitors

Answer 94

inhibitors that dont covalently change or modify the enzyme

Question 95

Reversible inhibition has 4 types:

Answer 95

Competitive Uncompetitive Non-competitive Mixed

Question 96

K_i

Answer 96

measure of inhibitor potency; the low this calue is, the more potent the inhibitor

Question 97

K_mapp

Answer 97

K_m in the presence of inhibitor

Question 98

V_maxapp

Answer 98

V_max in the presence of inhibitor

Question 99

Competitive Inhibitor

Answer 99

competes with substrate for the active site. addition of more substrate can overcome the effect of the inhibitor V_max stays the same K_M is increasing

Question 100

Non-competitive Inhibitor

Answer 100

binds to another site other than the active site that prevents the enzyme from doing its job well V_max \> V_maxapp K_M = K_Mapp

Question 101

Uncompetitive Inhibitor

Answer 101

binds to an enzyme already bound to a substrate that decreases the efficiency of the enzyme V_max \> V_maxapp K_M \> K_Mapp