Pre Knowledge (Chapters 2, 5, 12) Flashcards
(2) Proteins are polymers consisting of ..
amino acids
Proteins contain a wide range of … like alcohols, thiols, thioethers, carboxylic groups, amines and basic groups like amines
Functional groups
Components amino acid
central a carbon
R group (side chain)
amino group (NH2)
carboxyl group
Amino acids are chiral: explain
Mirror isomers: L isomer and D isomer.
Only L isomers are found in proteins
Zwitterions
Amino acids in solution at physiological pH have a dipolar form (NH3+ and COO-)
Hydrophobic / nonpolar amino acids
-Glycine, Gly, G
-Alanine, Ala, A
-Proline, Pro, P
- Valine, Val, V
- Leucine, Leu, L
- Isoleucine, Ile, I
- Methionine, Met, M
- Tryptophan, Trp, W
- Phenylalanine, Phe, F
polar uncharged amino acids
- Serine, Ser, S
- Threonine, Thr, T
- Tyrosine, Tyr, Y
- Asparagine, Asn, N
- Glutamine, Gln, Q
- Cysteine, Cys, C
- Histidine, His, H
Polar positive amino acids
- Lysine Lys, K
- Arginine, Arg, R
Polar, negative amino acids
- Aspartate, Asp, D
- Glutamate, Glu, E
Peptide bond
C=O - NH
Most common cross-ink within the linear polypeptide chain
Disulfide bonds
Most peptide bonds have the … configuration
trans
> Steric repulsion, two atoms closer tha Van der Waals contact distance > opposing force
The alpha helix is stabilized by intrachain …
hydrogen bonds
Characteristics of globular proteins: enzymes, signaling molecules, regulatory proteins
-Lack of symmetry
-Solubility in water
Which motif is found in many DNA binding proteins
Helix-turn-helix
(2) motifs
Secondary structures frequently present in proteins
(5) Catalytically active RNA molecules
Ribozymes
Properties of enzymes are speed and specificity. Which enzymes catalyze proteolysis of peptide bonds?
Proteases
The six major classes of enzymes
- Oxidoreductases: oxidation-reduction
- Transferases: group transfer
- Hydrolases: hydrolysis reactions (transfer of funcional groups of water)
- Lyases: addition or removal of groups to form double bonds
- Isomerases: isomerization (intramolecular group transfer)
- Ligases: ligation of two substrates at the expense of ATP hydrolysis
Ester bond
C=O
-O
Kinase and phosphatase function
Kinase: phosphorylation
Phosphatase: de phosphorylation
Molecules which regulate activity of the enzyme
Cofactor
Groups of cofactors
-Coenzymes (small organic molecules)
-Metals
Tightly bound coenzymes which are unchanged in the overall chemical reaction
Prostethic groups
> loosly associated coenzymes often behave like second substrates (cosubstrates) > bound by enzyme, changed and released.
Enzyme without cofactor
Apoenzyme
Catalytically active enzyme with cofactor
Holoenzyme
A reaction can take place spontaneously if …
dG < 0
A system is at equilibrium if dG =
0
Endergonic reactions
dG > 0, input of energy needed (or coupling to energetically favourable reaction(s)
Does dG give information about the rate of reaction?
No
dG in A + B <=> C + D
dG = dG’0 + RT ln([A][B]/[C][D])
How to measure dG0’
In equilibrium
dG0’ = -RT ln([A]eq[B]eq/[C]eq[D]eq)
because dG = 0
K’eq
= [A]eq[B]eq/[C]eq[D]eq
= e ^ -dG0’ / RT
RT = 2.48 at 298 K
pH = 7 in dG’0
Do enzymes alter the reaction rate? And the equilibrium?
Reaction rate: yes
Equilibrium: no
Equilibrium constants are the same with altered rate constants for foward and reverse reaction
What is the relative free energy of the transition state in a reaction?
Higher than that of subtrate and product.
Gibbs free energy difference between transition state and substrate
Activation energy.
An enzyme … the activation energy
lowers
Substrate-enzyme biding site mechanism
Induced fit
Free energy released during formation of large number of weak interactions between enzyme and substrate
Binding energy
Weak interaction
Ionic interactions, hydrogen bonds, Van der Waals interactions, Hydrophobic interactions/force
Km is the subtrate concentration at
1/2 * Vmax
A low Km means
High affinity of the enzyme for the substrate
Michaelis-Menten equation
v = Vmax * ([S]/[S] + Km)
Km equation
k-1+k2/k1
[E][S]/[ES]
Lineweaver-Burk plot characteristics
Slope: Km/Vmax
x- Intercept: -1/Km
y- Intercept: 1/Vmax
Axes
x: 1/[S]
y: 1/v0
Elasticity of the enzyme
The enzyme activity is sensitive to changes in environmental conditions (changes in [S])
- At [S] < Km: elastic but little activity
- At [S] > Km: active but inelastic.
Vmax and k-cat (k2) equation
Vmax = k-cat*[E]tot
[E]tot = [E] + [ES]
fraction of active site filled
f-es = V0/Vmax = [S]/([S]+Km)
Specificity constant
k-cat/Km
V0 = (k-cat/Km) [S][E]tot
Sequential reactions
All substrates must bind to the enzyme before any product is released.
Types of sequential reactions
-Ordered sequential: all substrates must bind in defined sequence.
-Random sequential
Double-displacement reactions
One or more products are released before all substrates bind the enzyme
Allosteric enzymes
Consist of multiple subunits and multiple active sites (binding sites), among one of them could be a allosteric actiation site
> no classic MM curve
Temperature increases enzyme activity until
denaturation temperature
Types of enzyme inhibitors
-Irreversible inhibitor
-Reversible inhibitors
Types of reversible enzyme inhibitors
-Competitive inhibition
-Uncompetitive inhibition
-Noncompetitive inhibition
Competitive inhibition
Inhibitor competes with the substrate for the enzyme
> has a higher affinity for the enzyme
> Only effective at low [S]
> If [S] increases, some [S] will bind nevertheless
How are Vmax and Km changed with competitive inhibition
Increase of Km
Vmax stays the same
Km after competitive inhibition
Ki (inhibitor) = [E][I]/[EI]
Uncompetitive inhibition
Inhibitor binds only to the ES complex
- Substrate dependent
- Prevents catalysis rather than substrate binding
- Cannot be overcome by increasing [S]
Km and Vmax after uncompetitive inhibition
Km lowered (Kmapp apparant Km)
Vmax lowered
Changes in Lineweaver Burk plot in UC inhibition
Same slopes, different x and y intercepts (higher since Km and Vmax lowers)
Noncompetitive inhibition
Inhibitor binds to different site than the substrate
- can bind free enzyme or ES complex, but decreases catalysis
- Cannot be overcome by increasing [S]
Mixed NC inhibition
Noncompetitive inhibition in which the inhibitor binds to its binding site preferably to either the free or bound state of the enzyme to the substrate
Km and Vmax after NC inhibition
Vmax lowered
Km not affected
What are potent competitive inhibitors?
Transition-state analogs
> resemble transition state
> bind tightly to active binding site
Irreversible inhibitors bind …
covalently
Groups of irreversible enzyme inhibitors
-Group-specific reagents
-Affinity labels
-Suicide inhibitors
group-specific reagents
inhibit enzymes by reacting with specific side chains of the enzyme
> not very specific
Affinity labels (reactive substrate analogs)
Are structurally similar to the substrate and covalently bind to active site residues (catalytic site)
> more specific
Suicide inhibitors
Also called mechanism based inhibitors
> use the mechanism of the enzyme to inhibit them, for example the enzyme changes the inhibitor into a reactive molecule which reacts with catalytic residues covalently.
- The enzyme commits suicide by generating its own inhibition
Molecular mechanism penicillin
Suicide inhibitor
- For the gram positive S aurus which has 1 membrane and a cell wall of peptidoglycans with crosslinks facilitated by glycopeptide transpeptidase (transfers peptide bond)
- penicillin has a beta-lactam ring which resembles the transition state of the transpeptidase reacion (R-D-ala-D-Ala) and has a peptide bond (reactive). And penicillin covalently binds the catalytic serine residue.
(12) Characteristics fatty acids
Long hydrocarbon chain with carboxylic acid groups
Saturated fatty acids have … double bonds
No
Lipid bilayer membranes contain carbohydrates bound to…
proteins or lipids
Membrane protein types
- Pumps
- Channels
- Receptors
- Energy transducers
- Enzymes
Membranes are …. assemblies (covalent/noncovalent)
noncovalent -> hydrophobic interactions
Membranes are (a)symmetric
asymmetric
Phospholipid components
-One or more fatty acids
-Platform to which fatty acids are attached (like glycerol)
-A phosphate
-An alcohol attached to the phosphate
How are phospholipids with a glycerol platform called?
Phosphoglycerides, bound to two FAs and a phosphorylated alcohol.
Bonds between glycerol and fatty acids
ester bonds formed by condensation of hydroxyl groups of glycerol and carboxyl groups of the FAs.
Sphingomyelin is a phospholipid in membranes derived from
a backbone of sphingosine (amino acohol with long unsaturated carbohydrate chain)
Second class of membrane lipids: glycolipids. Characteristics
- derived from sphingosine
- amino group bound to FA
- unit linked to the primary hydroxyl group is a glucose or galactose (instead of phosphorylcholine in sphingomyelin) with a ester linkage.
Third major type of membrane lipids beside phospholipids and glycolipids
Cholesterol
Structure cholesterol
-Four carbon rings sterol
- At one end: hydrocarbon tail
- other end: hydroxyl group which can interact with head group of phospholipids since it lays parallel to it in membrane
Membrane lipids important characteristic in solubility
Amphipathic: hydrophilic head group and hydrophobic tail group. –> fatty acids lay parallel to each other.
Micelle
globular structure formed from ionized fatty acids with single tails due to hydrophobic interactions in water.
Two opposing sheets of a lipid bilayer are called:
leaflets
lipid bilayers form …
spntaneously.
Forces in spontaneous bimolecular sheet formation
-Hydrophobic interactions
-Vanderwaals attractive forces favor tails laying close to each other
- Electrostatc and hydrogen bonding between polar head groups and water molecules
Lipid vesicles / liposomes
Aqueous compartments surrounded by a lipid bilayer
> can be synthesized artificially containing ions or molecules
Why does tryptophan, a zwitterion, transfer through a lipid bilayer (without transporter or channel) faster than K+ or Na+
It is amphipathic instead of solely hydrophilic
Myelin (membrane around neurons) contain many/little protein
little, it is a lipid rich membrane > form electrical insulator
Plasma membrane consist approximately …% of proteins
50%
Membranes with most proteins
mitochondria and chloroplasts (energy transduction membranes)
Classes of membrane proteins
- Integral membrane proteins: interact extensively with hydrocarbon chains of membrane lipids > only released by agents which compete with the nonpolar interactions
- Peripheral membrane proteins: bound to membrane with primarily electrostatic interactions and hydrogen bonds
Most common structural transmembrane motif
Membrane-spanning alpha helices
A beta barrel of beta strands can form a …
channel protein, alternating hydrophobic and hydrophilic amino acids along each beta strand
Prostaglandin H2 synthase is a integral membrane protein. How is it attached?
With a set of alpha helices with hydrophobic side chains on the outside of the surface to hydrophobically interact with the fatty acid tails
Which reaction is catalyzed by prostaglandin H2 synthase
Arachidonic acid (lipid) > prostaglandin G2 (by the cyclooxygenase (COX)) > prostaglandin H2 (by the peroxidase (POX))
How does aspirin lower inflammatory response, fever and pain
It covalently links its acetyl group (acetylsalicyclic acid) to a serine residue along the hydrophobic channel (with more hydrophobic interaction force than the lipid bilayer to catch the substrate btw) to block the channel irreversibly (no suicide inhibitor, not the catalytic residue)
> The COX cannot function and prostaglandin H2 is not synthesized, prostaglandins promote inflammatory response, including swelling, pain and fever.
How does ibuprofen inhibit prostaglandin H2 synthase? What type of inhibitor is it?
Ibuprofen is a competitive inhibitor which blocks the hydrophobic channel.
Lateral movement of membrane proteins and lipids
lateral diffusion
Lateral diffusion speed and transverse diffusion speed of lipids
Lateral diffusion: rapid
Transverse diffusion (flipflop): very slow (catalyzed by enzymes)
Membrane fluidity is dependent on
- Fatty acid composition
-Cholesterol content
Which FAs induce a more fluid membrane?
Unsaturated, less Van der Waals force.
Cholesterol makes the membrane more fluid/solid
solid, forms hydrogen bonds with carbonyl oxygen atom of phospholipid head group and hydrocarbon tail of cholesterol is located in nonpolar core of the bilayer
Do long or short FAs increase fluidity of a membrane
Small FAs: less Vanderwaals interactions
