Quiz 2 Flashcards
what happens with catalysis with multiple substrates
reactants brought together and properly oriented so they can react
glucose and glucokinase are an example of
the induced fit model for enzymes/substrates
what does glucokinase do
phosphorlyate glucose
when is glucokinase “open”
when no glucose is bound. active in closed state with bound glucose
what [S] is low, what is Vo proportional to
[S] – FIRST ORDER KINETICS
When [S] is high, what is Vo proportional to
equal to Vmax — 0 ORDER KINETICS
what does the rate depend on
the concentration of the substRATE
What does Km represent
the concentration of the substrate where Vo is 50% of Vmax
competitive inhibitor
binds at the same site
Km is increased but Vmax is the same, can reverse the effect by increasing the amount of substrate so that it takes up almost all the receptors on the enzyme
non-competitive inhibitors
doesn’t bind to same spot but makes enzyme less effective.
Km is the same, Vmax is decreased
uncompetitive inhibitor
binds ONLY when substrate is bound because ES complex creates a binding site
-Km AND Vmax are reduced
irreversible inhibitor
bind very strongly or produce covalent modification. Decreased Km but no change on Vmax – like NON competitive
Effector
binds non covalently to subunit of regulatory enzyme. Change in affinity or alters enzymatic activity in a positive or negative way
homotropic effector
substrate itself is the effector. THINK: Hemoglobin! Occupation of first site alters affinity of remaining
Heterotropic effector
substrate is not the effector
what kind of curve does a + cooperativity give
sigmoidal
positive heterotropic effector
binding at the regulatory unit of the enzyme causes conformational shift at the catalytic subunit. NOT COVALENT BINDING.
negative heterotropic effector
feedback inhibition – regulated enzyme typically catalyzes a rate limiting step `
what happens in calcium calmodulin path
Calcium binds to calmodulin cooperatively - with all 4 sites goes from “closed” to “open” and THIS is what binds to CAMKII
what happens when calmodulin binds CAMKII
conformational change which relieves auto-inhibition. CAMKII autophosphorylates so retains activity even after Ca returns to baseline.
What turns off calcium/calmodulin/CAMKII pathway
protein phosphatase which dephosphorylates CAMKII
zymogen
enzyme liberated by proteolysis of inactive precursor
where is trypsinogen released
by the pancreas into the duodenum
what cleaves trypsinogen to trypsin
enteropeptidase
cofactor
additional molecule that helps perform catalysis
metal ion or organic molecule which is usually referred to as coenzyme
apoenzyme vs holoenzyme
holoenzyme is protein + cofactor, apoenzyme is protein alone
isoenzyme
enzyme that catalyzes the same reaction
hexokinase
4 isozymes that phosphorylate simple sugars
glucokinase
hexokinase IV - this is the hexokinase in the liver. MUCH higher Km for glucose than others. Converts XS glucose to glycogen for storage
what is the advantage of the high Km of glucokinase for glucose
when glucose is scare, available glucose will be used by hexokinases in other tissues
Kcat
number of operations a single molecule of an enzyme can perform per second
specificity constant
how efficient enzyme is when free binding sites are available
what does a large specificity constant mean
rxn can proceed at high rate even if substrate concentration is low or enzyme is not highly expressed
how large can a specificity constant be and why
largest is 10^8 or 10^9 because limited by diffusion rates - enzymes that are at this rate are called “catalytically perfect”
what does heme effect
ALA synthase 7 steps earlier in heme synthetic pathway.
ALA is an allosteric enzyme whose activity is under control of an effector (heme).
what do we call inhibitors in pharmacodynamics
antagonists
what is the substrate in pharmacodynamics
drugs!
possible drug receptors
enzymes, channels, G protein coupled receptors
what gets measured with respect to drugs
amount of drug bound to receptor.. NOT rate of reaction
Vmax in pharmacodynamics referred to as
Bmax (max number binding sites)
agonist
drug with intrinsic effect on its target. EXCLUDES drugs that produce their effect by preventing some other ligand from binding because this isn’t intrinsic
EC50
on effect curve, concentration of drug that produces 50% of maximum effect of drug
Emax
on effect curve, area where max effect is produced
instrinsic efficacy
effect per molecule of agonist binding to its receptor
what happens when a signaling pathway is fully activated
this only happens with FULL agonists – even if we add more receptors, can’t produce larger effect!
will a partial agonist ever fully activate a pathway?
NO. Some receptors change conformation but downstream coupling doesn’t become as efficient so even with all receptors bound, mechanism won’t be FULLY activated
what is the relationship between Kd and EC50 for a partial agonist
Kd = EC50 (effect and binding curve are the same – each new molecule makes a difference)
what is the relationship between Kd and EC50 for a full agonist
KD > EC50 - Binding curve is to the R of the effect curve - Emax can be attained without all of the receptors being bound
what do full agonists often contain
spare receptors (ex: only need to activate 4 receptor sites for 100% activity)
drug potentcy
inverse of EC50
Inverse agonist
instrinsically decreases activity of a target protein BELOW its basal activity. This means the receptor must have some activity even without the drug present.
antagonist
no INTRINSIC effect on activity – produces effect by preventing endogenous ligands from exerting its effect. Analogous to inhibitors.
competitive antagonist
can be completely displaced from binding sites by high enough concentration of an agonist that binds to the same site
PA2
-log[agonist] that produces a 2 fold shift in EC50
what does it mean if you have a large PA2
low [antagonist] is enough to block agonist response
what does a high potency antagonist have
high PA2
therapeutic index
(toxic effect + TD50) / (therapeutic effect + ED50)
certain safety factor
lethal dose < D1 / therapeutic effect dose ED99
are drugs specific or selective for their targets
selective, NOT specific
how does a carrier protein work
utilizes ion gradients and works in cycles, binding solutes as they pass through the membrane
K+ charges in and out of cell, overall
Inside: 150
Outside: 4
Veq = -91
Na+ charges in and out of cell, overall
Inside: 10
Outside: 145
Veq: =67
Ca charges in and out of cell
Inside: 0.0001
Outside: 2
Veq: +125
Cl charges in and out of cell
Inside: 5
Outside: 110
equation for Veq
58/valence * log [So]/[Si]
Driving force
Vm - Veq
Ohm’s Law
V=IR or I = gV
why don’t real membranes exhibit instantaneous behaviour in response to applied current
because of capacitance – first charges the membrane, THEN can be used to create voltage difference
can carrier proteins move against gradient
YES, but channels cannot – they require ATP
Primary active transport
solute binds to pump and is directly carried through the membrane
Secondary active transport
pump establishes a large gradient for another solute, this gradient can be harnessed for energy
P class pumps
become phosphorylated over the course of a cycle
electrogenic
creates a current
ampipathic
hydrophobic and hydrophilic areas
SERCA
sarcoplasmic/ER Ca ATPase. Keeps Ca in cytoplasm low. High affinity binding site on cytoplasm of ER, binds Ca, phosphorylates, Ca transferred to lumen and sequestered
V/F Class pumps description
pump ONLY protons, contribute to acidification of organelles by pumping protons from cytoplasm into lumen of organelle
ABC Class pumps description and example
ATP Binding Cassettes – bind ATP at these conserved regions. Often transport uncharged or hydrophobic molecules. EX: Multidrug resistant Proteins and CF transmembrane regulator
Multidrug resistant protein
In epithelial cells, transport small, polar moelcules including some products of normal metabolism. Can also pump drugs out of cells, tumors that over express these are resistant to tx
CF transmembrane regulator
in lungs and other organs. Has no known “pumping” function. Has a channel permeable to Cl- and regulated by PKA. CF linked to mutation here so less Cl transport across pulmonary epithelial cells which leads to viscous mucous and bad gas exchange
Uniporter
conduct single cell species down gradient - help circumvent hydrophobic barrier (facilitated diffusion)
co-transporter
couple thermodynamically favored movement of one molecule with unfavorable movement of another.
1) Symporter: different solutes moved in SAME direction
2) Exchanger/antiporter: different solutes in opposite directions
GLUTs
bind one glucose - conformation changes, bring in and diffuses away, internal concentration of glucose kept low by phosphorylation of glucose
NCX
Na/Ca exchanger - couples Na IN to Ca OUT. Works with SERCA to regulate Ca.
Ratio: 3 Na for 1 Ca. Accumulates positive charge inside the cell. Used for muscle contraction in heart. Can slow calcium removal by interfering with pump! DIGOXIN!!
SGLTs
Na/Glucose transporters (Salt GLucose Transporters). Couple Na IN to glucose uptake. Kidney has 2.
1) SGLT 2 at early tubule: 1 Na for 1 glucose
2) SGLT 1 at late tubule: 2 Na for 1 glucose
what is the result of an excitatory stimulus
depolarized the membrane
what is the result of an inhibitory stimulus
hyperpolarizes the membrane by potassium conductance or activating Cl- conductance
what is the effect of opening Cl channels
often has minimal effect because reversal potential for Cl- is is close to resting membrane potential. BUT IT STILL DECREASES TOTAL MEMBRANE RESISTANCE!
what is the result of an excitatory current if Cl- channels are open, relatively speaking
reduces depolarization produces
Cys Loop channels examples
Nicotinic Acetylcholine Receptor (NaChR) and GABA-A receptor
Nictonic acetylcholine receptor
Mediates communication between neurons and muscles/neurons. Non selective cationic, depolarizes membrane (excitatory).
what is the target of myasthenia gravis
nicotinic acetylcholine receptor
GABA-A receptor
activated by gamma aminobutyric acid - main inhibitory receptor in the brain. Permeable to anions, inhibitory effect due to membrane hyperpolarization and decrease in total membrane resistance
glutamate receptors examples
AMPA or NMDA type
Cys loop description
5 protein subunits - have loop in extracellular N domain that is linked by cysteines. Each subunit has 4 membrane spanning helices -
M2 forms wall of channel
M3/4 has intracellular loop that associates with cytoskeletal partners
2 binding sites - BOTH must be activated
what passes through cys loop
cations or anions, not discriminating
NaChR description
Nicotinic acetylcholine - CYS LOOPS
- subunits are HIGHLY homologous
- all channels have at least two alpha subunits
- acetylcholine binding sites at alpha gamma and alpha
NaChR permeability and reversal potential
Permeable to Na and K but also to Mg and Ca. Reversal potential O mv - large enough to go through with hydration shells.
why is NaChR permeable to cations
negative residues lining outer regions of pore
how do NaChR channels go from being closed to open
unbound Ach - M2 helices have “kink”
bound Ach - M2 helices twist from center
GABA
CYS LOOP! Binds GABA. Mediates fast inhibitory transmission in the brain.
what is GABA permeable to and what is the Vrev
Cl- (anion selective)
Vrew: -80 mV
description of glutamate receptors
protein composed of tetramers, each subunit has three membrane spanning regions TM1, TM2, TM3. Pore due to loop in membrane from cytoplasmic linker between TM1 and TM2
AMPA function and Vrev
Selective for Na and K
Vrev = 0 mV
Binds glutamate
NMDA function and Vrev
Selective for Na, K and Ca.
Vrev : 20 mV
Blocked by extracellular Mg at resting Vm, expelled with membrane depolarization. Also requires glycine.
Inward rectifier description
formed from tetramers - 2 helical regions - M2 face each other in inverted teepee. P loop between M1 and M2 which is highly selective and has carbonyl groups
how does K pass through inward rectifier
after being dehydrated but gets stabilized by carbonyls (Na is too small to be stabilized, this is why it doesnt pass through)
how do KiR channels get blocked
positive charges on cytoplasmic side. Can’t get through selectivity barrier but can block. If membrane is depolarized, usually blocked.
when do voltage gated channels open
with depolarization
K+ voltage gated channels description
tetramers with TM helices S1-S6. S4 is charged and voltage sensitive. S5/S6 is what is inverted, S6 lines the pore.
deactivation
stay open until membrane repolarizes
inactivation
“ball and chain” - stop conducting current soon after activation, even if membrane stays depolarized. Basic AA at N terminus (ball and chain) can block the channel because the + charge is attracted to the pore
de-inactivation
membrane potential returns to normal so the inactivating particle leaves and goes to - cytoplasm. This process is NOT instantaneous
voltage gated Na/Ca channels
instead of four subunits have 1 polypeptide with 24 TM helices in 4 domains. homologous to seperate subunits of voltage gated K channels, selectivity filter is on the P loop. NO BALL AND CHAIN - inactivation from + charges on intracellular linkers
