Reading...
Play button
Play button
Use LEFT and RIGHT arrow keys to navigate between flashcards;
Use UP and DOWN arrow keys to flip the card;
H to show hint;
A reads text to speech;
81 Cards in this Set
- Front
- Back
|
RNAse protein class?
|
All-Beta strands
|
|
|
Which residue functions as an acid and which residue functions as a base in serine proteases?
|
Acid: Serine
Base: Histidie |
|
|
What metal ion does Carbonic Anhydrase use?
|
Zn 2+
complexed by 3 imidazole rings from Histidine |
|
|
What is the quaternary structure of Hemoglobin?
|
4 x myoglobin-like chains
all alpha helices D2 symmetry |
|
|
Which residues carry out concerted acid-base catalysis in RNAse A?
|
2 Histidines
His 12 = base, steals H+ from ribose's 2' hydroxyl His 119 = acid, donates H+ to leaving ribose |
|
|
Define Km, the Michaelis-Menten Constant.
|
Km is the substrate concentration at which Vo = 1/2 of Vmax.
|
|
|
What happens to the Lineweaver-Burke plot in competitive inhibition?
|
Slope changes.
Km appears to increase (competition for active site slows rate at which protein reaches 50% saturation) Vmax is same because catalytic ability is the same. |
|
|
What happens to the Lineweaver-Burke plot in UNcompetitive inhibition?
|
Y-intercept changes: Vmax decreases because catalytic ability is reduced.
Km appears to decrease because Vmax decreases as well. |
|
|
What is the function of Protein Disulfide Isomerase?
|
Repairs/catalyzes formation of disulfide bonds in peptide chains.
|
|
|
What is the single greatest contributor to protein stability?
|
The hydrophobic effect: entropic gain of water/solvent molecules. Protein folding hides hydrophobic residues inside globular protein.
|
|
|
What happens to the dG (Free Energy) of a protein as it nears its ideally folded conformation?
|
It decreases.
|
|
|
COLLAGEN
quaternary structure? composition? what are structures of subunits? |
Triple right handed a-helix
Repeating residues: Gly-Pro-Hyp Subunits: 3 left-handed helices wound around each other |
|
|
What holds collagen triple helix together?
|
Glycine N-H + Proline C=O hydrogen bonds between adjacent chains
|
|
|
Keratin:
How held together? |
A/D residues have hydrophobic interactions where the two alpha helices meet.
|
|
|
What are the two amino acids with chiral side chains?
|
Ile, Thr
|
|
|
What is the direction of twist in the keratin double-helix?
|
LEFT HANDED TWIST
|
|
|
Which segments of a peptide come off a gel chromatography column first?
|
Larger molecules, which cannot fit through the holes in the gel beads and pass through with the eluent.
|
|
|
What is the basis of the Edman Degradation?
|
It cleaves the N-terminal end of a polypeptide chain by reacting with phenyl isothiocyanate to create PTH derivative that is assayable once the peptide is subjected to acid hydrolysis.
|
|
|
What is the basis of Affinity Chromatography?
|
Column is packed with protein-specific ligand; impurities do not bind ligand and are flushed out. Protein of interest can then be cleaved from ligand to isolate.
|
|
|
What is the basis of protein dialysis?
|
Smaller proteins diffuse out through size-specific holes in the dialysis membrane; larger ones stay behind in the bag.
|
|
|
What is the basis of ion exchange chromatography?
|
Ion exchange chromatography separates proteins on the basis of how well they bind to a charged substrate at a given pH and salt concentration. Ex. if a protein has an esp. high net negative charge at pH = X, run it through a + column to separate it from a mix.
|
|
|
What is the basis of Sodium Dodecyl Sulfate Protein Agarose Gel Electrophoresis (SDS-PAGE)?
|
First SDS denatures proteins to linearize them. Then the large negative charge of SDS allows the gel to separate the pieces solely by size.
One SDS per any two residues --> charge proportional to size. Estimate molar mass. |
|
|
Where on hemoglobin does BPG bind?
What does it do? |
It binds the central cavity of deoxyhemoglobin.
It reduces hg's affinity for O2 by stabilizing the deoxy-hg state. |
|
|
What do areas of highly conserved protein sequence indicate?
|
A segment of protein that is important for proper function (i.e. active site or binding pocket)
|
|
|
What happens when the heme group binds Oxygen?
|
Fe2+ moves into the plane of the ring, pulling histidine and the a-helix with it to trigger the conformational change.
|
|
|
What is a sigmoidal curve indicative of?
|
Cooperative binding between protein subunits. Subunits must be in contact with one another and interact upon ligand binding.
|
|
|
What does the Bohr Effect have to do with Hemoglobin?
|
Ion pairs are disrupted in deoxyhemoglobin and get deprotonated in oxy state (Hb releases protons when it binds O2)
In acidic conditions these ion pair forming residues are protonated and stabilize/encourage hg to adopt deoxy state. High H+ --> lower O2 affinity |
|
|
Describe the SYMMETRY MODEL of allosterism.
|
Symmetry must be maintained (all or nothing change T/R states)
Ligand can bind either T or R state Binding encourages shift to R state. |
|
|
Describe the SEQUENTIAL MODEL of allosterism.
|
Binding of subunit necessitates conformational change.
Symmetry need not be maintained Binding in one increases binding in neighbors through slight structural changes. |
|
|
What is the protein class of calmodulin?
|
All-alpha
Two domains Undergoes conformational change upon ligand binding. |
|
|
What is the critical flaw in sickle cell hemoglobin, HbS?
|
Glu6 --> Val6
Val fits in a hydrophobic pocket in other HbS or Hb and makes them stick together. |
|
|
What is the best buffer range for weak acids and bases?
|
pK +/- 1 pH point
|
|
|
What is the definition of Kcat?
|
The ratio of product produced per unit of enzyme. Kcat = K2 ish.
A measure of enzyme's efficiency. |
|
|
What is alpha in the context of enzyme kinetics?
|
in competitive inhibition, it is the factor by which [S] must increase to overcome impact of [I]
|
|
|
What is the mechanism of neuraminidase?
|
Neuraminidase = influenza protein.
Cleaves sialic acid from the membrane of host cell to allow new virus particles to escape. |
|
|
What is a Michaelis Complex?
|
ES, the enzyme/bound substrate complex.
|
|
|
What is the x-intercept on the Lineweaver-Burke Plot?
|
1/Km
|
|
|
What is the y-intercept on the Lineweaver-Burke Plot?
|
1/Vmax
|
|
|
Define "prochiral."
|
An achiral molecule that can be made chiral in one step (one switch, etc).
Ex. Citrate --> Isocitrate |
|
|
What is the structure and class of neuraminidase?
|
4 domains of all wiggly beta strands
|
|
|
What are the final two proteins involved in the blood coagulation cascade?
|
Prothrombin --> Thrombin
activates Fibrinogen --> Fibrin |
|
|
What are two ways the body regulates enzyme activity?
|
1. make inhibitors
2. make proenzymes and activate as necessary. |
|
|
What makes DANA a suitable inhibitor?
|
It is a neuraminidase transition state analog
Planar like transition state of sialic acid 4-guanidino group makes ion pair with glutamates in active site --> high affinity for neuraminidase. |
|
|
Is it better for an enzyme to bind the substrate or the transition state tightly?
|
Transition state. Stabilizing the transition state lowers the activation energy of the reaction.
|
|
|
In the serine proteases, which residue acts as a base and which acts as an acid?
|
His is a base taking H+ from Ser (acid)
His is an acid donating H+ to leaving peptide His is a base abstracting H+ from H2O, then an acid donating to Ser (base). |
|
|
What is a prosthetic group?
|
An organic or inorganic coenzyme that is covalently attached to the enzyme.
|
|
|
What is the Reaction Coordinate?
|
The reaction path of lowest free energy.
|
|
|
Why is the peptide bond planar?
|
Resonance between the N lone pair and the carbonyl double bond gives the C-N peptide bond partial double bond character.
|
|
|
Which side chains are aliphatic?
|
Gly, Ala, Val, Ile, Leu, Pro
|
|
|
Which side chains are aromatic?
|
Phe, Tyr, Trp
|
|
|
Which side chains are polar and uncharged?
|
Met, Ser, Thr, Cys, Asn, Gln
|
|
|
Which side chains are negatively charged (acidic)?
|
Asp, Glu
|
|
|
Which side chains are positively charged (basic)?
|
Arg, Lys, His
|
|
|
What is the structure and class of myoglobin?
|
All alpha
Eight helices Monomeric Globular unit, Soluble |
|
|
What is the structure and class of ATCase?
|
C6R6 (Catalytic, Regulatory)
D3 symmetry mixed a/b domains |
|
|
What is actin made up of?
|
Repeating G-Actin subunits arranged in helical symmetry
|
|
|
What molecules inhibit ATCase and how?
|
CTP through feedback inhibition (allosteric modification)
PALA through competitive inhibition (bisubstrate analog) |
|
|
What molecule activates/stimulates ATCase?
|
ATP through allosteric modification
ATP binds ATCase away from the active site. |
|
|
What enzyme activates chymotrypsinogen?
|
Trypsin.
Later pi-chymotrypsin activates itself and makes alpha-chymotrypsin. |
|
|
Which three endopeptidases (of the six we studied) are serine proteases?
|
Trypsin
Chymotrypsin Elastase |
|
|
What is the mechanism of RNAse A?
|
Two Histidines at active site --> concerted acid/base catalysis to form cyclic ribose phosptate intermediate
|
|
|
What is the difference between a cis and trans peptide?
|
Orientation of side chains w/respect to peptide bond.
Cis = on same side Trans = on opposite sides (usually more stable) |
|
|
What two important biological molecules are derived from tyrosine?
|
Dopamine
Thyroxine |
|
|
What is the name for aggregates of misfolded proteins?
|
amyloids
Alzheimer's, Transthyretin diseases, Mad Cow Disease (prions) |
|
|
What are the alpha-keratin helix specs?
handedness, length, pitch, etc. |
Right handed helix
450 Angstroms long 310 residues long 3.5 residues/turn 5.1 Angstrom pitch |
|
|
What was significant about Adenosine and GAPDH?
|
Hydrophobic substituent on Adenosine fit nicely into groove on sleeping sickness parasite GAPDH enzyme.
Inhibitor drug design |
|
|
Class and structure of Trypsin?
|
All beta protein
Monomeric unit Serine Protease |
|
|
What three methods of covalent catalysis are utilized by serine proteases?
|
1. concerted acid/base catalysis
2. transition state stabilization (oxyanion hole) 3. covalent catalysis |
|
|
How are proteases more frequently controlled?
|
Synthesis of a protease inhibitor molecule (ex. PTI, Pancreatic Trypsin Inhibitor)
|
|
|
What family of enzymes does Thrombin belong to?
|
Serine Proteases
|
|
|
Which conformation of ATCase is favored by binding of CTP?
|
Closed "T" conformation, less active.
|
|
|
PALA binds the active sites of ATCase, causing a conformational change from T --> active R.
Why does it still inhibit catalysis? |
It doesn't let go of the active site. So even though ATCase is in active state it can't catalyze reactions.
|
|
|
How many molecules of sialic acid can neuraminidase bind?
|
Four: one per wiggly all-beta subunit.
|
|
|
What is the symmetry of GroES?
|
C7
|
|
|
What is the symmetry of GroEL?
|
D7
|
|
|
Where do side chains point in a beta sheet?
|
Alternating above/below the plane of the sheet.
|
|
|
Where do side chains point in an alpha helix?
|
Toward the outside of the helix.
|
|
|
Where are active sites most frequently found?
|
At the junctions between adjacent protein domains.
|
|
|
How much more stable is folded vs unfolded protein?
|
dG = ~0.4 kJ/residue/mol.
100-residue protein: only ~40kJ/mol more stable |
|
|
Describe the three-step renaturation process.
|
1. Expose to oxygen to reoxidize (reform) scrambled disulfide bonds.
2. Remove Urea to allow folding. (still scrambled) 3. Add catalytic amount of beta-mercapthoethanol to repair scrambled disulfide bonds --> renatured protein. |
|
|
What is the significance of the Hill Coefficient?
|
Steepness of Hemoglobin Binding Curve
Steeper Hill Coefficient --> faster change in Hg O2 saturation/unloading. |
