Examination 1

Front 1

Biomolecules

Back 1

1) Protein
2) Lipids
3) Carbohydrates
4) Nucleic Acids

Front 2

Amino Acids
Monosaccharides
Nucleotides

Back 2

Proteins
Carbohydrates
Nucleic Acids

Front 3

CHON

Back 3

Makes up 96% of human body weight. Nitrogen, Phosphorus, Sulfur and CHON are the 6 most important elements in the body.

CHO is 98% of atoms in an enzyme.

Front 4

Proteins

Back 4

Most versatile biomolecule. Protein function depends on structure. Enzymes, signaling, receptors.

Structural support, transport, signaling, receptors and enzymes.

Front 5

Nucleic Acids

Back 5

DNA and RNA. Made of nucleotides. Nucleotides had a phosphate group (for energy), a pentose sugar and a nitrogen base. Its function is to store information.

Carry ATP, signaling molecules.

Front 6

Lipids

Back 6

Used as energy storage and as barriers. Smaller then proteins and nucleic acids. Can be phospholipids (main membrane component), Fats/Triacylglycerides (energy storage) or steroids/sterols (structural support and signaling). Hydrophilic head and hydrophobic tail.

Front 7

Phospholipids

Back 7

Have glycerol and fatty acids. Also have a phosphate group and a variable group (triglycerides do not). 2 fatty acid chains.

Front 8

Triglycerides

Back 8

Have glycerol and fatty acids. 3 fatty acid chains

Front 9

Fatty Acids

Back 9

Used to make triglycerides. 6 times more energy than sugars by weight. Also used to make phospholipids.

Front 10

Carbohydrates

Back 10

Sugars. Functions as an energy source, energy storage, information molecule and sometimes as structural support. Glucose is stored in animals as glycogen. Cell walls in plants (cellulose). Linked to proteins (glycoproteins to proteoglycans.

Front 11

Central Dogma

Back 11

DNA to RNA is transcription. RNA to protein is translation. DNA polymerase catalyzes replication. RNA polymerase catalyzes translation.

Front 12

Organelles

Back 12

h

Front 13

Receptors

Back 13

Are stereospecific. NH3 is to the left means that the amino acid is "L". "L" is preferred.

Front 14

Thermodynamics

Back 14

Non-spontaneous reactions can proceed through by coupling. ATP is the energy currency of the cell.

Front 15

1st Law of Thermodynamics

Back 15

Energy can't be created or destroyed. Energy of the universe is constant, but can be transferred.

Front 16

2nd Law of Thermodynamics

Back 16

Law of increased entropy (disorder) Natural processes tend towards equilibrium (disorder).

Front 17

Thermodynamic Values

Back 17

"H"= Enthalpy. Total heat content. Forming or breaking of bonds
"S"= Entropy (disorder).
"G"= Gibbs free energy. Total energy available in the system.
"DeltaG"= Free Energy change.

Front 18

When DeltaG is Negative

Back 18

Exergonic, spontaneous. Energy is released.

Front 19

When DeltaG is Positive

Back 19

Endergonic. Non-spontaneous. Energy must be used.

Front 20

Gibbs Free Energy Equations

Back 20

DeltaG= DeltaGfinal-DeltaGinitial

DeltaG=Change in Enthalpy-T(change in entropy)

Front 21

Water

Back 21

Most abundant substance in cells. Water forms hydrogen bonds. They can hydrogen bonds with other water molecules and other substances in the cell. Water molecules are polar because the individual bonds are polar and the molecule is asymmetrical.

Front 22

Weak Interactions (Non-covalent Interactions)

Back 22

Don't involve electron sharing. Based on charge attraction and the hydrophobic effect.

1) Hydrogen Bonding
2) Ionic (Electrostatic)
3) Van der Waal's
4) Hydrophobic Effect

Front 23

Hydrogen Bond

Back 23

Between H on one molecule and an O molecule on another. Gives water a higher melting point, boiling point, heat of vaporization and surface tension.

H-O and N-H

Front 24

Ionic

Back 24

Salt bridge (Amino to carboxyl) formation in proteins. Ionic interactions are greater in non-polar environments. Water has a high dielectric constant (D, ability to store electrical energy in an electric field) compared to non-polar solvents.

Front 25

Van der Waal's

Back 25

Asymmetry in electrical charge. Not a bond, more of a force of attraction.

Front 26

Hydrophobic Effect

Back 26

Hydrophobic molecules cluster together. Increases entropy. In amphipathic molecules, water will interact with the hydrophilic ends.

Front 27

The Hydrophobic Effect

Back 27

The hydrophobic effect is the observed tendency of non-polar substances to aggregate in aqueous solution and exclude water molecules.

Front 28

pH

Back 28

pH= -log (H+)

pH of pure water is 7. As [H+] increases, pH decreases.

Water can ionize.

As "H" increases, pH decreases.

Front 29

Weak Acids

Back 29

Acetic acid. Bicarbonate and Hydrogen phosphate.

Front 30

Henderson-Hasselbach Equation

Back 30

pH= pKa+log[A+]/[HA].

Front 31

pKa

Back 31

The pH at which acid is half dissociated. pH above the pKa means that there is more A-, pH below the pKa means that there is more HA.

When the pH=pKa, 50% ionization occurs.

Front 32

Weak Acids

Back 32

R-CH2-COO-
R-CH2-NH2
HCO3-
HPO4

Front 33

Strong Acids vs Weak Acids

Back 33

In a strong acid, most of the molecules break up into solutions. In a weak acid, fewer molecules break up into ions.

Front 34

Buffers

Back 34

1) Acidosis: Shift of -.2pH
2) Alkalosis: Shift of +.2pH

Front 35

Hemoglobin

Back 35

As pH goes up, affinity goes up. Can be protonated of N-terminally modified (carbamate) to transport H+ or CO2. Blood pH is 7.5, +/-.5 pH.

Front 36

Amino Acid Structures

Back 36

Have 4 different groups connected to the tetrahedral alpha-carbon atom. Amino acids are chiral. Only "L" amino acids are the constituents of proteins.

Front 37

Negative (Acidic) Amino Acids

Back 37

1) Aspartate (Asp, D)
2) Glutamate (Glu, E)

Front 38

Positive (Basic) Amino Acids

Back 38

1) Lysine (Lys, K)
2) Arginine (Arg, R)
3) Histidine (His, H). Aromatic.

Front 39

Polar Amino Acids

Back 39

1) Serine (Ser, S)
2) Tyrosine (Tyr, Y). Aromatic
3) Threonine (Thr, T)
4) Cysteine (Cys, C)
5) Asparaginine (Asn, N)
6) Glutamine (Gln, Q)

Front 40

Hydrophobic (Uncharged) Amino Acids

Back 40

1) Glycine (Gly, G)
2) Alanine (Ala, A)
3) Valine (Val, V)
4) Proline (Pro, P). "R" group bonds to chiral carbon.
5) Tryptophan (Trp, W). Aromatic.
6) Leucine (Leu, L)
7) Isoleucine (Ile, I)
8) Methionine (Met, M)
9) Phenylalanine (Phe, F). Aromatic.

Front 41

Essential Amino Acids

Back 41

1) Histidine
2) Isoleucine
3) Leucine
4) Lysine
5) Methionine
6) Phenylalanine
7) Threonine
8) Tryptophan
9) Valine

Front 42

Zwitterions

Back 42

Dipolar ion. Charge on amino acids at physiological pH.

As the pH rises, COOH is the first to give up a proton.

Low pH, both groups are protonated.
Physiological pH, both groups are charged.
High pH, charge on carboxyl and no charge on amino.

Front 43

Functional Roles of Proteins

Back 43

They have a role in dynamic and structural functions.

Front 44

Dynamic Functions

Back 44

Storage and transport (Hemoglobin for O2 and ferretin for Iron).

Myosin and actin allow for contractions and changes in cell shape.

Catalysis of metabolic reactions (enzymes).

Front 45

Structural Functions

Back 45

Connective tissue and cytoskeleton.

Front 46

Protective Functions

Back 46

Immunoglobin and interferon.

Fibrin stops blood loss (helps blood clotting).

Front 47

Control Functions

Back 47

Hormones allow control and changes. Insulin (sugar homeostasis) and somatotropin (growth hormone).

Control and regulation of gene expression (Transcription factors, polymerases, histones).

Front 48

Protein Structure

Back 48

Peptide bond formed through a condensation reaction and is strongly covalent. Alpha of carboxyl to alpha of amino. Loss of 1 water molecule due to condensation.

1) Fibrous Proteins: Long and thread-like. Serve as structural components (provides strength)

2) Globular Proteins: Compact shapes. Serve as enzymes and other functional proteins.

Front 49

Primary Protein Structure

Back 49

Amino acid sequence. Covalent bonds.

Proline or Glycine often found at bends or turns in polypeptide chain.

Front 50

Secondary Protein Structure

Back 50

Alpha helices and beta sheets. Local folds. Hydrogen bonds between the backbone (not the side chains).

Front 51

Helix Breakers

Back 51

Proline and glycine. Glycine has conformational stability. Proline has a cyclic structure and no hydrogen from its amide to hydrogen bond.

Front 52

Alpha Helices

Back 52

Held by hydrogen bonds between every 5th amino acid. Side chains are on the outside of the helix. 3.6 amino acids per turn. Right-handed helix is the correct form because there is less steric hindrance.

Front 53

Beta Sheets

Back 53

Flat. Side chains project below and above the plane of the sheet. H-bonding between backbone atoms hold the sheets together.

Front 54

Alpha Helices and Beta Sheets

Back 54

Arise through H-bonds between N-H and C=O groups of the peptide bonds. Side chains are not involved.

Front 55

Tertiary Protein Structure

Back 55

Many different domains. Hydrogen bonds, electrostatic interactions, hydrophobic interactions, covalent (disulfide) bonds, ionic interactions and Van der Waal forces.

Large globular proteins have many different domains that have different motifs. proteins with similar functions have similar structural motifs.

Front 56

Domains

Back 56

Any structure in polypeptide that can independently fold into its own secondary and tertiary structure. Often have a specific purpose.

Front 57

Quaternary Protein Structure

Back 57

Not all have it. Hemoglobin does.

Front 58

Fibrous Proteins

Back 58

1) Alpha Keratin: Intermediate filaments (cytoskeleton). 2 right-handed helixes twist together. Left-handed superhelix. Contact between helixes by hydrophobic residues. Quaternary stabilized by disulfide bonds.

2) Collagen: Component of connective tissue. Glycine-Proline-Hydroxyproline. Each chain is a helix. Helix formed due to steric repulsion or proline residues, not H-bonds. 3 left-handed twists in a right-handed fashion (collagen helix). 35% glycine, 11% alanine, 21% proline/4-hydroxyproline.

Front 59

Osteogenesis Imperfecta

Back 59

Other amino acids are present instead of glycine. Collagen folds wrong, accumulates and causes cell death. Results in fragile bonds.

Front 60

Scurvy

Back 60

No vitamin C (ascorbate) so collagen defect. Vitamin C needed to make hydroxyproline. Hydroxyproline needed for stable collagen. Results in skin lesions, tooth loss, fragile blood vessels,

Front 61

Transmissible Spongifrom Encephalopathies

Back 61

Prions. Misfolded proteins. Mad-Cow, Creutzfeldt-Jakob and Scrapie (sheeps).

Alzheimers, Parkinson, Huntington and spongiform lead to protein aggregates which leads to cell death.

Front 62

Hydrophobic Effect

Back 62

Entropically-driven folding because of shielding of hydrophobic residues.

Front 63

Chaperones that Assist Folding

Back 63

1) Protein Disulfide Isomerase. Help coordinate disulfide bonds.

2) Heat-Shock Proteins: Help stabilize partially unfolded proteins.

3) Help block hydrophobic forces of proteins to stop aggragation.

Front 64

The Proteome

Back 64

All of the proteins expressed in a cell.

Front 65

Chromatography

Back 65

1) Gel Filtration/Size Exclusion Chromatography

2) Ion Exchange Chromatography: Charged beads.

3) Affinity Chromatography: Receptors on beads.

Front 66

Immunoblot/Western Blot

Back 66

Detection of protein with fluorescent antibody emission.

Front 67

Enzyme-Linked Immunosorbent Assay (ELISA)

Back 67

Antibody added to antigen. Substrate added and converted by enzyme into colored product.

Front 68

Isoelectric Focusing

Back 68

Separate based on charge (pH).

Isoelectric point (pI): Net charge on protein is zero
No SDS

Front 69

Polyacrylamide Gel Electrophoresis (PAGE). 2D Gel.

Back 69

Proteins denatured by SDS. Reducing agent can break sulfide bonds, not SDS. Protein separates on size.

Front 70

Amino Acid Analysis (AAA)

Back 70

A peptide or protein undergoes acid hydrolysis
6 M HCl in sealed, vial at high temperature (>100°C)
The constituent amino acids are separated by chromatography. Amino acid separation done by Ion Exchange HPLC. Provides amino acid composition, not the order in the chain.

Trp, Cys, Ser and Thr can degrade under acid hydrolysis.

Asn and Gln can undergo deamination.

Front 71

Edman Degradation

Back 71

Phenyl isothiocyanate reacts with N-term aa.
Breaks aa off & can be identified by chromatography

Front 72

Centrifugation

Back 72

1) Differential Centrifugation: Separate certain organelles from whole cells.

2) Ultracentrifugation: In an analytical ultracentrifuge, a sample being spun can be monitored in real time through an optical detection system, using ultraviolet light absorption and/or interference optical refractive index

3) Gradient Centrifugation

Front 73

Antibodies

Back 73

Immunological separation: Antibodies.

Front 74

Enzymes

Back 74

Catalysts, specific an allow transition states to form. Enhance the reaction rate. Enzymes are very selective. Alter reaction rates, not the reaction equilibrium.

Front 75

Enzyme Specificity

Back 75

Proteases cleave (hydrolysis of peptide bonds) at specific regions along a peptide bond.

1) Trypsin: Cleave C-terminal side of Arg and Lys.

2) Thrombin: Cleaves Arg-Gly bonds.

3) Aminopeptidase: General cleavage of N-terminal side.

4) Chymotrypsin: Aromatic amino acid residue cleavage.

Front 76

Enzymatic Terms

Back 76

1) Active/Substrate Binding Site: 3D site of an enzyme where catalysis takes place.

2) Cofactor: Vitamin derivative needed in many enzyme reactions, includes metals.

3) Apoenzymes: Enzymes with cofactors.

4) Holoenzymes: Enzymes with cofactor bound.

5) Isozyme: Different enzyme from different genes with similar sequences that catalyze the same reaction.

Front 77

Activation Energy (Gibbs Free Energy of Activation)

Back 77

Determines reaction spontaneity, it is a thermodynamic parameter, not a kinetic one and is independent of pathway. Energy required to start reaction is ΔG*. ΔG* is free energy of activation energy. Enzymes affect this, not the DeltaG.

Front 78

DeltaG

Back 78

ΔG = ΔG° + RT ln([B]/[A])

Front 79

Transition State

Back 79

High-energy conformation. Bridges initial state (reactant) to the final state (product).

Front 80

Enzyme/Substrate Models

Back 80

1) Lock and Key: Active site is complementary to the substrate.

2) Induced Fit: Enzyme forms shape that is complementary to the substrate upon substrate binding.

Front 81

Keq

Back 81

Keq>1, ΔG° is negative
Keq<1, ΔG° is positive

Front 82

Keq and ΔG°

Back 82

ΔG°= -RTln(Keq)

LogX =Y means 10Y=X
lnX = Y means eY=X
lnX = 2.3logX
Log 1 = 0
Ln 1 = 0
If a number is > 1, the log and ln are both positive
If a number is < 1, the log and ln are both negative

Front 83

6 Enzyme Classes

Back 83

1) Oxidoreductases: Oxidation is electron loss, reduction is gain. Electrons moved form 1 substrate to another like hydride ion or H.

2) Transferases

3) Hydrolyases: Add -OH from water to substrate (proteases)

4) Lyases: Catalyse double bond formation through double bonds.

5) Isomerases: Move groups or double bonds in a molecule to yield isomeric forms (rearrangement)

6) Ligases: Make C-C, C-O and C-N bonds. Needs ATP. Pyruvate carboxylase adds CO2 to pyruvate to make oxaoacetate, needs coenzyme biotin and ATP.

Front 84

Transferases

Back 84

Transfer a chemical group from 1 molecule to another. 2 substrates, 2 products.

1) Hexokinases: Transfer phosphate from ATP to glucose.

2) Kinases: Transfer phosphate groups from NTPs.

3) Aminotransferases: Move amino groups. Convert keto acids to amino acids. Requires pyridoxal phosphate (Vitamin B6) as a coenzyme

Front 85

Coenzymes

Back 85

Small organic molecules often from vitamins. Metal ions are cofactors. Can be modified during the reaction, but restored after. Stabilizes charges. Don't bind at the active site, don't participate in the reaction.

Front 86

NAD and NADP

Back 86

Coenzymes. NAD made form niacin (B6)

NADH= Catabolic (ETC)
NADPH= Anabolic

Front 87

FMN and FAD

Back 87

Both derived from riboflavin (B2)

Prosthetic groups tightly/covalently bound to enzyme.

Do oxidoreduction reactions. FMNH2 and FADH2

Front 88

Pyridoxal Phosphate (B6)

Back 88

Amino acid related reaction. Transamination. Modified and restored during the reaction by the enzyme.

Front 89

ATP

Back 89

Provides energy from terminal phosphate. Allosteric modulator (binds to regulatory site, not the active site and causes a conformation change).

Hydrolysis of ATP favorable due to energy release caused by the - charges on ATP through steric hindrance.

Front 90

Metals

Back 90

Mg2+ used in ATP reactions and associates with APT, not the enzyme.

Alcohol Dehydrogenase: Zn
Superoxide Dismutase: Cu and Zn
Heme: Fe
Pyruvate Kinase: K

Front 91

Mechanisms of Catalysis

Back 91

1) Covalent Catalysis: Enzyme forms transient covalent bond with the substrate.

2) Acid-Base Catalysis: Acid or base speeds up the reaction while not being consumed. Amino acid function groups accept or donate proton.

3) Metal-Ion Catalysis: Charge stabilization, oxidation-reduction.

Front 92

"V"

Back 92

Veloctiy (Rate of catalysis in mol/s)

Front 93

Velocity

Back 93

Rate at which substrate disappears (or P appears)/time.

Related to substrate [ ] via rate constant (k).

V=k [A]

Front 94

"Vmax"

Back 94

Max Reaction Velocity.

All enzyme has substrate bound. So at Vmax, ES=[E]t

Vmax= k2[E]t

At high [S], limiting factor is [E].

Front 95

"Vo"

Back 95

Initial rate. Product concentration is low.

Defined as increase in product when "P" is low.

Vmax/2

Front 96

Michaelis Equation

Back 96

Vo= Vmax*[S]/[S]+Km

Front 97

Km

Back 97

Michaelis constant. Km= [S] at which 1/2(Vmax). Estimates binding strength (substrate's affinity for an enzyme).

Can approximate [S] in the cell.

If Km is small, low [S] needed to fill active site. ES complex is strong (high substrate affinity)

If Km is large, high [S] needed, ES complex is weak.

Front 98

Turnover Number

Back 98

Turnover rate of enzyme (# of S molecules converted to P molecules per unit time when enzyme is saturated). # of P molecules formed per second at Vmax.

k2=kcat=turnover rate. k2=Vmax/[E]t

[S] is usually at, or below the Km so enzymes are usually not saturated with substrate.

Front 99

Catalytic Efficiency

Back 99

Kcat (rate of enzyme catalysis with substrate)/Km

Front 100

Multiple Reactants Reactions

Back 100

1) Sequential Reactions: All reactants bind to enzyme before product release.

2) Double Displacement (Ping Pong) Reactions: Product released before all reactants are bound to enzyme.

Front 101

Proteins Function in Groups

Back 101

1) As Large Complexes: Many enzymes in 1 large multi-protein complexes.

2) Linear Pathway: Sequential metabolism of a substrate.

Front 102

2 Levels of Regulation

Back 102

1) Genetic: Gene turned off, enzyme is not made.

2) Stop/Promote binding.

Front 103

Enzyme Regulation

Back 103

Most common mechanism involves feedback.

Allosteric Enzymes: Regulated by molecules binding to other sites (not the active site).

Front 104

Commitment Step

Back 104

Where the reaction goes through or not.

Front 105

Feedback Inhibition

Back 105

Involves control by the amount of product formed, reducing the activity of the enzyme in the pathway.

Positive Feedback: Product feeds back to increase its own production. Negative regulation is more common.

Positive Regulation: Increased activity through binding by ligand. Allosteric enzymes.

Front 106

Allosteric Enzymes

Back 106

Don't follow Michaelis-Menten kinetics.

Front 107

Concerted Model (Allosteric)

Back 107

4 assumptions

1) Allosteric enzymes have multiple subunits/active sites.

2) R=relaxed state/active conformation.
S=tense state/more stable, less active.

3) T/R ration= allosteric constant (Lo)

4) Symmetry Rule: All subunits are either T or all R.

Front 108

T and R

Back 108

Substrate binds more to R form than to T form.

Front 109

Cooperativity

Back 109

Binding of S disrupts T-R equilibrium in favor of R.

Front 110

Threshold Effect

Back 110

After threshold [S] reached, enzyme activity increases.

Front 111

Sequential Model

Back 111

Sequential changes in subunit structure. Conformational change in 1 subunit induces change in others= Affinity for substrate increases.

Addition or removal of phosphoryl groups from a protein is the most commons modification to affect protein function.

Ser, Thr and Tyr have PO3. Kinase adds PO3. Phosphatase removes it. Phosphorylation increases or decreases a protein's activity.

Front 112

Factors that Alter Enzyme Activity

Back 112

1) Temperature: Increase in reaction followed by denaturation.

2) pH: Increase in reaction followed by denaturation. Changes the charge on protein.

Front 113

Competitive Inhibition

Back 113

Reversible with more [S]. Inhibitor same in structure as the substrate. Sulfanamide and PABA are enzyme inhibitors.

Increase in [I] leads to Km increase, no effect on Vmax.

Front 114

Uncompetitive Inhibition

Back 114

ES to ESI. No product formed. Decrease in Vmax. Decrease in Km. Reversible

Front 115

Noncompetitive Inhibition

Back 115

S and I bind simultaneously to enzyme at different sites. Decrease in # of active enzyme molecules. If they don't bind at the same time, product is still formed. Decrease in Vmax (ESI makes no product), unchanged Km. Reversible

Front 116

Enzyme Regulation

Back 116

Not all enzymes are regulated, only important ones.

Front 117

Irreversible Inhibitors

Back 117

Substrate mimics that can't come off the catalytic residue. Increase [S] does not help. Enzyme active site is blocked.

1) Group-Specific Reagents: React with R-groups of amino acids. In active site.

2) Affinity Labels/Substrate Analogs: More specific for active site. TPCK binds at chymotrypsin active site and modifies histidine residue. Inhibitor looks like substrate.

3) Suicide Inhibitors: Mechanism-based. Inhibitor processed catalytically by enzyme, forms intermediate that modifies enzyme and inactivates it. Penicillin.

4) Transtion-State Analogs: Protein Inhibitors. Inhibitor mimics transition-state substrate, binds enzyme thus making it unavailable for the substrate.

Front 118

Chymotrypsin

Back 118

Chops at aromatic ring. Also a digestive enzyme. Likes phenylalanine.