Biomacromolecules, Protein Structure Flashcards
Name the 4 macromolecules
Carbohydrates
Nucleic acids
Proteins
Lipids (but these are not covalently bonded)
What are sugars
Straight chain polyhydroxy alcohols, including an aldehyde or ketone group
What is a glucose molecule
What is its usual form
6 C sugar with an aldehyde group on C1
Cyclic
What different forms of glucose are formed when it becomes cyclic
Why
α glucose
β glucose
New chiral centres are formed
How is β glucose arranged
Chair conformation
NOT planar but puckered
What is fructose, how is it different to glucose and what ring does it form
A 6 C sugar with a ketone group on position 2
5 sided ring
What is the bind formed why two monosaccharides are linked by a condensation reaction
Glycosidic bond
Which end of a sugar chain is the reducing end
Where the ring can be opened to produce a free reducing group (the aldehyde end)
Name 2 common disaccharides
Lactose
Sucrose
What is starch a polymer of
What is starch used for
α glucose
To store energy in plants
What is glycogen a polymer of
What is glycogen used for
α glucose
To store energy in animals
Are glycogen and starch branched?
Yes, but starch is more tightly packed so glycogen has more free ends from which glucose can be cut
What is cellulose
Unbranched chain of glucose connected by β linkages
V strong due to H bonds
Only termites can break it down in their digestive tract
What are proteins and lipids coated in
Complex carbohydrates (oligosaccharides)
What is RNA
A working template involved in gene expression and an information store in some viruses
It has a structural role in some cellular machinery eg ribosomes
Describe the sugar in DNA
Where is it deoxy
Deoxyribose is an aldopentose whose β-anomer is used in DNA
C2
What is the base used in DNA
Purines or pyrimidines with extra groups
What are the purine bases
A
G
What’s re the pyrimidine bases
C
T
U
Give the complementary base pairs
A-T
G-C
A-U
How is the stranding different in RNA to DNA
RNA is single stranded
How is the double helix arranged
How often is a complete turn
2 poly-nucleotide strands arranged with an external phosphate backbone and bases pointed to the centre like ladder rungs
Every 3.4nm
Why does DNA have major and minor grooves
Provide access to the bases for DNA binding proteins
Why does a hairpin loop form in RNA
Some regions are complementary to other regions in same strand
How does the HIV virus use RNA
It is its genetic material containing many loops and hairpin structures, which is important to pack the RNA into the virus capsid and time control gene expression
What is a ribosome molecularly
What are the active sites made of
A complex of RNA and protein
It is a ribozyme- an RNA molecule that acts as an enzyme
RNA
Give the 4 levels of protein structure
Amino acid sequence
Local backbone arrangement
3D fold
Arrangement of subunits
What do all amino acids contain?
An amino group, a carboxyl group and a H around a central α C with a 4th R group
What form does the α carbon take in proteins
L form
Which amino acid has no side chain
Glycine
Give an amino acid with an aliphatic and hydrophobic side chain
Alanine (R group is just methyl)
Why is proline special
R group forms a ring of 3 CH2’s, linking to the N
It can therefore not form H bonds
Give an amino acid with
a) a negatively charged R group
b) a positively charged R group
a) Arginine
b) aspartic acid
Which amino acid has a polar and aromatic R group
Tyrosine (with -OH coming off the aromatic ring)
What part of asparagine is polar
The NH2
Give an aromatic R group amino acid
Phenylalanine
Which amino acids contain sulphur
Cysteine
Methionine
Why are Cys residues unique
-SH groups can form a disulphide bond, a covalent crosslink that stabilises protein structures
How are peptide bonds planar
e- delocalisation
There is free rotation only around the α carbon
How does folding of a polypeptide occur
Rotation of φ (phi) and ψ (Psi)angles
Which is the N terminus and which is the C terminus
How Is an amino acid sequenced
N= has a free NH3+ C= has a free COO-
Sequence is numbered from N to C
What was the first protein whose primary structure was determined and when
How are they now determined
Insulin’s structure was determined chemically in 1955
DNA sequencing
Describe the experiment used to show that primary structure provides all the information needed for tertiary structure
- Pure RNase was dissociated by adding urea and mercaptoethanol
- Urea disrupts non-covalent forces holding the protein together
- mercaptoethanol reduces disulphide bonds
- when these denaturing agents were removed, protein spontaneously refolded
How are secondary structures formed
What are the 2 main types of secondary structure
How are side chain interactions involved
H bonds between N-H and C=O groups
α helix and β sheet
They are not
Which amino acid doesn’t have an NH
Proline
When are H bonds strongest
When linear
What does the Ramachandran Plot show
The combination of φ and ψ angles found in proteins
Shaded regions are combinations found in secondary structures
Any other combination of angles leads to steric clashes
Describe the α helix structure
Backbone in the middle with side chains pointing outwards
Right handed
H bonds are between C=O of residue i and NH of residue i +4
2 types of β sheet
Anti parallel - strands go in opposite directions
Parallel - same direction ( residue makes H bonds with residue 2 and 4 of other sheet)
Side chains always point alternatively above and below the sheet
How are a) parallel and b) anti parallel sheets joined
a) long loop of protein
b) short loop forming a hairpin
Give the 5 major interactions that form tertiary structure
H bonds Hydrophobic effect Electrostatic interaction VdW forces Disulphide bonds
Describe the hydrophobic effect
Water molecules cannot form H bonds with hydrophobic molecules
Water molecules become ordered around hydrophobic side chains, which is entropically unfavourable
To avoid this, hydrophobic molecules cluster together and interact with each other
Where are the hydrophobic and hydrophilic side chains usually on a protein
Hydrophobic in the core
Hydrophilic outside/ on surface
Give another name for electrostatic interactions
Salt bridges
How does water affect electrostatic interactions
Water molecules shield charges so interactions are stronger inside a protein as there is no water there
Describe disulphide bridges in proteins
Covalent bonds between 2 cystine side chains
Do not form inside the cell as it is a reducing environment
What kind of disulphide bonds do intracellular proteins form
They do not form disulphide bonds
How strong is the hydrophobic effect
Between VdW and H bonds
Give the structure of fibrous proteins and give 2 examples
Regular, ordered, with a strong repeating unit
Collagen
Keratin
Give structure of collagen
Is the collagen helix an α helix
Each chain forms a helix which twists around 2 others to form a right handed super helix 🧬
No it only has 3 residues and it is left handed (goes anti clockwise)
What is each of the 3 collagen chains made up of
Copies of a 3 amino acid repeat: Gly-X(usually proline)- Y(usually hydroxyproline)
What gives collagen its strength
The tightly packed triple helix structure
How are the 3 amino acids arranged in collagen
Proline on the outside of the triple helix
Gly On the inside, with its small size allowing 3 helices to pack closely
Proline hydroxylation allows another H bond to form
What are motifs
Name some and describe how they are held together
Commonly observed groupings of secondary structure elements
β-α-β: H bonds between strands and hydrophobic interactions between helix and strands
α-helical hairpin: hydrophobic and ionic
Greek key: H bonds between strands
What is a domain
Part of the protein which forms a structurally Independant unit with a hydrophobic core
Often composed of several motifs put together
It is a structural and functional unit
How many polypeptide chains and domains do antibody molecules contain
4 polypeptide chains
12 domains
What do B lymphocytes do
Produce antibodies
What is clonal expansion
When an antibody recognises an antigen, the cell producing that antibody is stimulates to replicate and secrete more
Describe an antibody molecule
Tetramer
2 identical light chains and 2 identical heavy chains
Stabilised by disulphide bonds
Each chain has a variable at the N terminus which confer binding specificity
Describe the domains in light and heavy chains in antibody molecules
Light: one variable VL, one constant CL
Heavy: one variable VH, 3 constant HC
How do the different domains in antibody chains interact
VL and VH associate to be complementary to the binding site/ the epitope of the antigen
CH1 and CL associate
CH2 and 3 drive dimerisation and interact with different receptors
What are cofactors
Molecules used by proteins to provide chemical reactivity not found in amino acid side chains
Define prosthetic group
It is tightly bound to the protein
What is a co-substrate
Loosely attached and used once then released after reaction for regeneration
For NAD+, give the:
a) vitamin source
b) metabolic role
c) reaction role
a) Niacin
b) Redox reactions involving 2 e- transfer
c) Cosubstrate
For FAD, give the:
a) vitamin source
b) metabolic role
c) reaction role
Riboflavin
Redox for 1 or 2 e- transfer
Prosthetic group
For CoA, give the:
a) vitamin source
b) metabolic role
c) reaction role
Pantothenate
Acyl group transfer
Cosubstrate
For Tetrahydrofolate, give the:
a) vitamin source
b) metabolic role
c) reaction role
Folic acid
One carbon substituent transfer
Provides methyl group for T in DNA
Co- substrate
For TPP, give the:
a) vitamin source
b) metabolic role
c) reaction role
Thiamine
Aldehyde transfer
Prosthetic group
How are NAD and FAD reduced/ oxidised
Which is endergonic
Accept/ release 2 e-
Reduction is endergonic
Oxidation is exergonic
What does NAD+ accept
2 e- and a H+
Give an example where NADH is used for reduction
Which molecule is assayed one this experiment and why
Pyruvate to lactate catalysed by lactate dehydrogenase
[NADH] - only molecule that absorbed light with a wavelength of 340nm
How does FAD become FADH2
Accepts 2 e- and a H+ onto each double bonded N
What is haem
Prosthetic group used to bind to O molecules
Where does the O bind on a haem unit
Central Fe which is always Fe2+ in Oxygen carriers
Why can haem also carry e-
So Fe can be reduced to Fe2+ or Fe3+
How many amino acids and helices does myoglobin contain
What does it do
153 amino acids
8 helices
Facilitate diffusion of O2 in muscle tissue
Describe the graphs of saturation vs pO2 for myoglobin and haemoglobin
What does this show
Myoglobin- hyperbolic
Haemoglobin- sigmoidal
Cooperativity
What is the Fe in haem bound to
4 N in the porphyrin ring and 1 N on the His side chain
How does haemoglobin change shape when it becomes oxygenated
In deoxy- it is done shaped with the Fe above the ring
O2 binding pulls it into the ring’s plane, pulling down the His with it
This causes changes in the whole tetramer and shifts other subunits closer to the R state, making it easier for O2 to bind
Is T state oxy or deoxy
Deoxy
How do ion channels work
Allow hydrophilic ions to cross membranes through a pore
True or false: the K channel is a dimer
False it is a tetramer if identical helices
How does the selectivity filter work in a K channel
It strips the aqueous shell from K ions and provides C=O O atoms that allows the loss of hydration shell without the loss of energy
Na is too small to contact C=O so it stays hydrated and is too large
How can you determine protein structure
X Ray crystallography
NMR
How does x Ray crystallography work
Ordered molecule crystals distract x Rays and an electron density map is calculated
What is cryoEM
Cyroelectron Microscopy
Many images are taken and averaged to generate a single image
Describe affinity chromatography
The column material contains a molecule that specifically binds to the protein of interest. When the sample passes through only that protein will bind and others wash away.
The protein can then be eluted from the column, usually by adding a competitive ligand
Describe ion exchange chromatography
Uses a column of charges materials
Proteins will bind to different degrees depending on their charge
Proteins are eluted with increasing salt which disrupts electrostatic attractions
Highly charged proteins elute at higher salt concentrations
Describe gel filtration chromatography
Column contains a gel of porous beads. Small proteins can pass through the pours into the beads, while larger proteins pass around them.
Therefore small proteins take longer to pass through
What is another name for gel filtration chromatography
Size exclusion chromatography
What diseases can be caused by misfolding
Parkinson’s
Alzheimer’s
CJD
BSE
What are the brain plaques in Alzheimer’s disease patients surrounded by
dead neurons
Amyloid fibres of a misfolded protein Aβ are found in these dead cells
What is protein Aβ
Aβ is a degradation product of amyloid precursor protein
What test shows that proteins are the infectious agent in Alzheimer’s
What causes Aβ to misfold
Injection of amyloid fibrils into animals leads to development of disease
Unknown
What are prion diseases such as kuru caused by
Misfolding of the PrP protein -
PrP^c is normal, PrP^sc is misfolded Scrapie form
Misfolding is catalysed by PrP^sc, leading to infectivity
How much protein?
Use the Bradford assay
Use a standard/ calibration curve of [protein] vs absorbance
When would you use SDS PAGE
To determine which protein
How does SDS PAGE work
DeNature protein by heating and SDS binds to hydrophobic regions of protein to make it negatively charged (it needs to be charged) so you know which way it will run
You compare the marks with proteins of known mass
Describe ELISA
Antibody 1 is on plate
Add sample with >2 domains
Bind antibody 2 to sample to check it has stuck
Antibody 2 is covalently bonded to an enzyme
When a substrate is added, the enzyme converts into a product that can be detected
Done in 96 well plate
What is used in HIV testing
ELISA
Reactions catalysed by enzymes show what properties
How do enzymes drive energetically unfavourable reactions
Increases rates
Great reaction specificity
Capacity for regulation
Haha TRICK QUESTION enzymes cannot drive energetically unfavourable reactions
How can enzymes provide a driving force for reactions
Coupling a favourable to the unfavourable one
What is a more realistic model of enzyme function
Induced fit
Where both enzyme and substrate change conformation when they interact
Describe a case study for the induced fit model
Carbonic anhydrase
Active site of carbonic anhydrase contains a zinc ion coordinated with 3 His side chains and a water in 4th position
Binding to zinc deforms water, polarising the e- and His64 accepts this e-
Why are HIV proteases important for HIV
Why is this a important for treatment
HIV makes its molecules as ‘polyproteins’ which must be cleaved by HIV protease (an Asp protease) to be functional
Human cells do not do this cleavage so the protease is an excellent drug target
How are inhibitors designed generally
To block to enzyme’s active site and binding of the protein substrate
How were HIV protease inhibitors developed
Designed to be similar to substrate and to mimic tetrahedral transition state of cleavage reaction. However they cannot be cleaved.
Other chemical groups were also added to improve solubility and stability of drug
Name a HIV drug
Have such drugs been useful?
Are there any problems?
Saquinavir
HIV drugs have led to a 70% reduction in AIDS deaths in areas where they are available
Virus can develop resistance through mutation of residues in drug-binding pocket
Give the 6 classes of enzymes according to IUBMB
Oxioreductases Transferases Hydrolases Lyases Isomerases Ligases
Give the function and an example of oxioreductases
Catalyse oxidation and reduction reactions (ie removal and addition of e-)
Alcohol dehydrogenase removes hydrogen from alcohol
Give the function and an example of transferases
Transfer a functional group from a donor to acceptor molecule
Protein kinases transfer a phosphate group onto a protein molecule
Give the function and an example of hydrolases
Catalyse hydrolytic cleavage
Eg HIV protease or DNase (which cuts DNA
Give the function and an example of lyases
Break (covalent) bonds by means other than hydrolysis or oxidation, creating new double bonds or rings
Aldolase yin glycolysis which breaks fructose-1.6-bisphosphate and generate a C=O bond
Give the function and an example of isomerases
Catalyse geometric changes
Proline racemase
Triose phosphate isomerase in glycolysis
Give the function and an example of ligases
Join molecules together using ATP
DNA ligase which joins 5’ end of one DNA molecule to the 3’ end of another
Give the order of glycolysis
Glucose➡️ glucose-6-phosphate ➡️ fructose-6-phosphate ➡️ fructose-1,6-bisphosphate
⬇️⬇️
GADP↔️DHAP
Where do you find glucokinase and hexokinase
What are the different properties
Glucokinase = liver
Hexokinase - muscle
When blood glucose is low:
Glucokinase has low activity so does not uptake glucose
Hexokinase: active so muscles continue to use glucose
Give the Michaelis-Menten equation for enzyme to product
Why is ES —> E+ P irreversible
E+S 🔁 ES ➡️ E + P
Under initial conditions, [P]=0 and reverse reaction is therefore minimal
What is the actual Michaelis Menten Equation
Vo = Vmax[S]
—————
Km + [S]
What is Vmax and Km
Which axis is each on
Vmax= maximum rate at this enzyme concentration
Y axis
Km= substrate concentration where rate is Vmax/2
X axis
What happens if you put Vo= Vmax/2
[S] = Km
What is the steady state assumption for michaelis menten
[ES] is constant
Ie rate of formation=rate of breakdown
Vmax=?
K2x[E]t
2 requirements for mechalis menten equation
Only [S] is changed when Vo is measured
[E]«
When is Vmax achieved
What is the equation for Vo here? Therefore what is a simple way to increase Vmax
When [S] is infinite
There is no free enzyme, it is all within the ES complex
Vo=k2[E]total= Vmax
Increase [E] by making more enzyme
What is the equation for the catalytic constant when Vmax is achieved
Why
Vmax
———-
[E] total
K2=Kcat
What is the turnover number
Number of reactions per second at active site
In relation the the Michaelis Menten graph and ES complexes, what happens as the [substrate] increases
The percentage of enzyme in the ES complex increases and the rate increases towards Vmax
Give a simple operational definition of Km
The substrate concentration at which the enzyme works at half of its maximum rate
What is Kd and what does it measure
K1/K-1 (ie the rate of E+S —>ES divides by the rate of ES—-> E+S)
The affinity of E and S
What is the formal definition of Km
K-1+K2
———-
K1
K1= E+S—-> ES K2= ES—->E+P K-1= ES—-> E+S
An enzyme with a low Km has a ____ affinity
High
Give the units of Km
M or moldm-3
What is the Lineweaver Burk plot
If we measure the initial rate for a series of different substrate concentrations and plot 1/Vo against 1/[S] we will get a straight line
Give the rearranged equation of the Michaelis Menten equation as
y=Mx+c
Y= 1/Vo
M=Km/Vmax
X=1/[S]
C=1/Vmax
How do competitive inhibitors work
Bind to active site of enzyme, blocking access for the substrate
Binding is transient ie inhibition is reversible
What is the best competitive inhibitor
One that resembles the transition state
Talk about the inhibition of proline racemase
It is an enzyme that interconverts L-proline and D-proline with a planar transition stage
Inhibitors mimic the transition state and bind with 160x the affinity than proline (the substrate)
How does Ritonavir work
It is a competitive inhibitor of HIV protease designed to mimic its transition state
Why does a competitive inhibitor increase Km without affecting Vmax
More substrate is needed to reach Vmax/2
How does a non-competitive inhibitor affect Vmax and Km
Explain
Changes Vmax but not Km
It binds to both E and ES
It does not affect substrate binding but reduces catalytic activity
How do irreversible inhibitors work
They form covalent bonds with essential active site residues, preventing substrate entry
Describe the cell wall of bacteria, it’s enzyme and the effect of penicillin
Bacteria are protected by a peptidoglycan cell wall of cross linked sugars and peptides
Glycopeptide transpeptidase cross links peptidoglycan chains during cell wall synthesis but is inhibited by penicillin so the bacteria continues to grow but the cell wall doesn’t form correctly and bursts due to osmotic pressure
How does penicillin inhibit glycopeptide transpeptidase
It is a suicide inhibitor
Penicillin binds to the O on glycopeptide transpeptidase’s Ser and the reaction cannot be completed
Talk about Sarin gas
It inhibits acetylcholine esterase at the NMJ
Irreversible inhibitor
Deadly
Why does sarin bond more readily to acetylcholine esterase
Acetylcholine as there is a
HO-C=O
(making the C δ positive for the enzyme to bind to)
Sarin has a
F-P=O
In the same place so the more electronegative F is lost more easily
When is reversible covalent modification useful and what is a common example
Enzyme control
Phosphorylation
Which residues can phosphate groups be added to
The -OH of Ser, Thr, and Tyr
Give the structure of a phosphate group
O-
|
O=P-O-
|
O
|How is cyclin dependent kinase 2 controlled
Phosphate is inserted onto Tyr preventing binding of ATP
This must be removed for activation
How can phosphorylation activate an enzyme
Give the example of zymogens
Allosteric changes
Zymogens are generated in an inactive form and are activated by proteolytic cleavage
Which state is Oxy for haemoglobin
Oxy= R Deoxy= T
Give an example of a monomeric allosteric enzyme
Glucokinase (found in pancreatic β cells)
It has a binding cleft between two domains which reorientate when the substrate binds (from super-open (inactive) to closed (active))
In the presence of glucose an intermediate state is slowly formed but then this quickly becomes closed when ATP is present
What forms of Diabetes is glucokinase associated with
MODY and noenatal diabetes (if the enzyme is mutated)
How big is an angstrom
10^-10m
What does the allosteric site do In glucokinase
Stabilises active state
How do glucokinase activators work as drugs
Reduce the Km (so there is a higher binding affinity) and therefore increase glucose sensitivity thus increases insulin secretion
Why do you not want to affect the Vmax of glucokinase activators
The enzymes don’t work faster overall, causing hypoglycaemia
What kind of curve would an allosteric enzyme have
Why
Sigmoidal
All enzymes initially in T state
Eventually enough are in R state for sudden increase in rate (when there is a very high affinity)
Why can you not use Michaelis Menten for allosteric enzymes
Can you talk about Vmax
No single Km is always correct
Yes - it will reach a maximum rate
Where does an enzyme want to be on a [S] vs [activity] graph
At the steep bit so a small change in [S] causes a large change In Activity
What does an activator do
Ditto for inhibitor
Discuss their respective graphs
Stabilises the high activity form (curve becomes hyperbolic)
Stabilises low activity form (still sigmoidal but higher [S] is required to switch to high activity form)
What does PFK do
Binds fructose-6-phosphate and ATP
Does ATP inhibit PFK
why
Yes
If ATP levels are high you do not need energy so glycolysis is inhibited
Give the structure of PFK
4 subunits with each monomer contributing to the active site and allosteric sites at each subunit interface
ATP inhibits by being to allosteric site
ATP can never be v low in cells so how can PFK be regulated
Give experimental proof
AMP
Even the addition of 0.1mM of AMP to 1mM of ATP pushes the sigmoidal curve to the left into a hyperbolic curve
Give the equation for the reaction that glycogen phosphorylase controls
Glycogen(n residues)+phosphate ——> glucose-1-phosphate + glycogen (n-1 residues)
How is glycogen phosphorylase controlled
ATP and glucose-6-phosphate inhibit
AMP activates
Phosphorylation of Ser-14 stabilises high activity form (stabilising phosphorylase a form)
(a for active)
What must a successful drug have (3)
High selectivity High affinity High bioavailability (stays in the patient)
What must an influenza virus do to infect a cell
Haemagglutinin binds to Salic acid on the host cell surface and then get inside to hijack cell’s machinery
The neurominidase is necessary to release new virus particles as it cuts the Salic acid off the surface so the new virus doesn’t get stuck
How were influenza drugs designed
What was the problem
To mimic Sialic acid transition state - it had 1000x more affinity
Poor selectivity so attacked human cells
How was the influenza drug improved
Chemical groups were added including a positively charged group to form hydrogen bonds with 2 glutamate side chains
Further, parts of the molecule that didn’t interact with the enzyme were removed to make it more lipophilic and therefore could pass through membranes (improving bioavailability)
Why do we think bird flu is resistant to Tamiflu
There is a large cavity in the neuraminidase
What is the story of Viagra
Initially developed as angina treatment (a planned phosphodiesterase block)
But after trial ended, test subjects did not want to give back the drug
Was this discovered to have an effect on erectile dysfunction
What was the idea behind viagra
If you inhibit breakdown of cGMP, it should lead to vasodilation
cGMP leads to activation of PKG which leads to vasodilation
Eventually it was changed so that Viagra is now basically an analogue of the substrate (cGMP)
What are the characteristic angles associated with α helices
Dihedral angles
Which molecules decrease enzyme activity by increasing Km without changing Vmax
Competitive
How many carbon atoms in citric acid
6
Which gene is most strongly associated with a predisposition to cancer
APC
Which DSB repair is vital in G0 and G1
NHEJ
Which DNA repair stalled replication forks
HR
What is the initial step in mRNA degradation in eukaryotes
Deadenylation
How do you convert nucleotides in RNA editing reactions
Hydrolytic deamination
What are a category of genetic markers comprising repeats of 1-5 nucleotides in length
Micro satellites
What causes Prader Willi syndrome
A deletion of the paternal 15q11b
Commonly associated with eating disorders and short stature
What causes VFCS
Non allelic recombination between related repetitive sequences in chromosome 22
How do enzymes catalyse a reaction
Stabilising the transition state
How do serine proteases work
They use a reactive serine residue to perform nucleophilic attack on the substrate molecule
Which B vitamin are vegans likely to be deficient in
Vitamin B12 (cobalamin)
What is a physiological activator of guanylyl Cyclase
NO
What are riboswitches
RNA sequences that directly interact with small molecules to control translation
True or false
Eukaryotic PIC is defines as a set of specific DNA binding transcription factors bound to an enhancer element
False
How does the glucocorticoid receptor bind to promoters
As a dimer
What does AKT
Phosphorylates the target of mTOR protein
What is the action of β catenin in the Wnt pathway
It trans locates from the cytoplasm to the nucleus
What increases due to increased IP3
Ca2+
What substance increases inside the cell due to generation of nitric oxide
cGMP
Which infectious protease contains 2 essential Asp residues in its active site
HIV protease
Name an enzyme inhibited by penicillin
How is it inhibited
Glycopeptide transpeptidase
Suicide inhibition
Which enzyme uses a catalytic Zn ion for transition state stabilisation
Carbonic anhydrase
Which is the only amino acid that can form a covalent bond with another amino acid
Cysteine (because it has a -SH group that can link to another -SH group in a disulphide Bridge )
Which amino group can’t form H bonds
Proline
Describe the reaction mechanism of serine proteases
1) Ser performs nucleophilic attack on C in substrate
2) His accepts the new Ser H+
3) Asp stabilises positive charge on His. The tetrahedral intermediate is now formed
4) tetrahedral decomposes, accepting His H+, and the first product is formed and is replaced by H2O. The Ser is now separate, bound to acyl-enzyme
5) His accepts H+ from H2O and OH in the water attacks C=O of acyl enzyme. This forms a second tetrahedral which decomposes to release the enzyme
What are the roles of the catalytic triad
What is the oxyanion hole
This is important in serine proteases
Ser- nucleophilic attack
His- acid/base
Asp- stabilises charge on His
The hole is only filled when the transition state is formed on the way to making the tetrahedral intermediate
What does the active site of carbonic anhydrase contain (3)
How does the reaction work
Zn coordinated by 3 His side chains.
The H2O substrate occupies the 4th coordination position
Active site also binds CO2
Binding to Zn deforms H20 so His can take a H+
The remaining OH- attacks the C of CO2, generating HCO3-, which is released
What kind of enzyme is HIV protease
How do these enzymes work
Asp protease
Use Aspartic acid as reactive groups
There are 2 Asp: 1 in an environment favourable for protonation (A) and 1 in aq environment (B)
H2O is coordinated between them
B removes a H+, allowing the water to nucleophilic attack the C=O at the peptide bond
The CN bond is broken and the N pulls B’s H+ towards it and the products are released
How many molecules of Acetyl CoA does 1 palmitoyl CoA
8
How many e- are involved in Vit C redox reactions
2
Where are the H bonds on an α helix
Between C=O of i and NH of i+4
What are α helical hairpins held together by
Mostly hydrophobic (and some ionic) interactions
