biochem lectures 6 & 7 pt 2 Flashcards
what is ATP synthase
multisubunit transmembrane protein
another name for ATP synthase
complex V
what is ATP synthase made of
2 subunits, F1 and F0
what is F0
water insoluble transmembrane proton pore
why is F0 water insoluble complex
cuz the parts of the transmembrane proton pore that forms interacts w/ inner mitochondrial membrane, so there’s a lot of interactions w/ hydrophobic fatty acid side chains
what does complex IV do
uses energy of reduction of O2 (to H2O) to pump one H+ into the intermembrane space for each e- that passes through
how much NADH to reduce one O2
2 NADH
how much H+ is used to reduce O2
4 H+
is F0 subunit free floating or embedded
it’s embedded within inner mitochondrial membrane
what pore structures do protons diffuse thru
this F0 subunit of ATP synthase
basically what is F0
pore structure, what protons will diffuse thru
what is F1
catalytic component
describe F1
water soluble peripheral membrane protein complex
what carries out the actual synthesis of ATP
F1
how does ATP synthase work
by coupling facilitated diffusion of protons thru F0 w/ catalytic mechanism underlying synthesis of ATP via F1 component
how much ATP does ATP synthase generate
1 ATP per every 3 protons
what happens to F0 as proteins diffuse thru it
begins to rotate
where does rotation of F0 occur
within inner mitochondrial membrane
what does rotation of F0 complex affect
gamma protein
what is gamma protein
a part of F1; physically connecting thru other accessory proteins
what happens as F0 turns
causes gamma protein subunit to turn with it
what happens as gamma subunit turns
causes conformational changes to alpha and beta subunits in F1 catalytic complex
describe alpha-beta units
3 sets of alpha-beta dimers or pairs in F1; each is capable of synthesizing ATP
what is rotational catalysis or binding chain mechanism
rotation of F0 drives movement of central gamma chain, gamma chain tweaks alpha-beta pairs of F1 that helps drive conformational change in catalytic subunits needed to make ATP
what is F0 again?
transmembrane proton pore; part of ATP synthase thru which protons will diffuse through
what does this pore structure enable
enables protons to, via facilitated diffusion and PMF, to pass thru F0
what is atomic force microgaphy
type of electron microscope, allows you to look at C subunits in F0
what are C subunits
form pore structure of F0
what is F1
catalytic subunit of ATP synthase
describe structure of F1
alternating a-b pairs/dimers around a central gamma subunit
where is gamma subunit located
in center of F1
how many sets of alpha-beta pairs or dimers in F1
3 sets
at any given time, what can each of those alpha-beta pairs assume
1 of 3 conformations; b-ATP, b-ADP, b-empty
what does each conformation represent
a diff step or ability/function of those alpha beta pairs to do something relevant to ATP synthesis
b-ADP
not catalytically active
what does b-ADP bind
ADP and Pi
b-ATP
catalytically active
what does b-ATP bind
ATP
what is b-empty
low affinity for ATP or ADP
which conformation has a high affinity of alpha beta pairs for ADP and Pi
beta-ADP
what is catalytically active form
beta-ATP
what form of alpha-beta do we get ATP synthesis
beta-ATP
what is beta-empty
as it transitions from beta-ATP to beta empty, we get release of ATP thats just been formed
what happens as central gamma subunit turns
causes each alpha-beta pair to go thru b-ADP, b-ATP, and then beta empty conformation
what is order of sequence of conformations
b-ADP, b-ATP, beta-empty
at any give point, describe each of the alpha-beta dimers
each is in one of those three states or conformations
why do we call it beta-whatever
because its beta subunit that is important for catalysis
what is alpha subunit needed for
functionality
what is rotational catalysis
free E generated w/ proton movement is harnessed to interconvert the conformation states to make and release ATP
what are the conformational changes driven by
rotation of rotor relative to aB subunits
how much H+ used for each 120* turn
each 120* turn uses 3 H+
conversion of what is related to synthesis of ATP
beta-ADP –> beta-ATP
conversion of what is related to release of ATP
b-ATP to b-empty
how does beta ADP transition to beta ATP conformatoin
as gamma subunit rotates (cuz F0 rotates), brings about conformational changes within three sets of alpha-beta pairs
what results in movement of central gamma subunit
diffusion of protons thru F0, causes rotational catalysis to occur, results in movement of central gamma subunit
what do we have at any given time
we will have each of these alpha-beta pairs in one of these 3 conformational change
how much does it rotate
120* increments
what happens w/ each 120* turn of subunit
brings about conformational change in each of the three alpha-beta pairs
what is each rotation driven by
thru diffusion of protons thru F0
what happens to an alpha-beta pair that was in b-ADP conformation after a 120* rotation of gamma subunit
converted to b-ATP conformation
what happens after another 3 protons diffuse thru (after b-ATP)
beta-empty conformation
what happens after it releases ATP
resets
what happens when one b-subunit assumes b-empty
one neighbor assumes b-ADP, one assumes b-ATP
what does one complete rotation of gamma cause
causes each b to assume all 3 conformations
what happens per 360* turn
produces 3 ATP
who came up w a way to visualize rotational catalysis
japanese scientists; used recombinant proteins tagged w/ histidine, and nickel coated glass
what else did japanese scientists notice
120* increments
yield of cell respiration
3 ATP per NADH, 2 ATP per FADH2
what do we get from anaerobic fermentation
only 2 ATP per glucose
total yield per glucose for cell respiration
30/32
why do we utilize more PE in glucose thru cell respiration
because we synthesize greater numbers of ATP in cell respiration vs. fermentation rxns
what is efficiency of ATP production from PE stored in glucose
30-40%
how much of PE stored in glucose are we losing, and to what
60%, in form of heat
what gives us body temperature
metabolic activities of body
how much PE from glucose is released in form of heat
greater than 50%
what is 40% harnessed to make ATP sufficient for
enough for necessary biochemical rxns; we need to consider additional E sources (fats, proteins, etc)
what is ox phos coupled to
electron transport
what happens if you block electron transfer
block ATP synthesis
what do you need in order for ATP synthesis
electron flow; source of electrons
can we uncouple ATP synthesis from electron transport
yup
what drugs can uncouple this
oligomycin or venturicidin
what are oligomycin and venturicidin
ATP synthase inhibitors; enzymes that block ATP synthase’s ability to do what it does
what happens if you treat ATP synthase with these ddrugs
not only fails to allow diffusion of protons thru F0, but prevents synthesis of ATP
what happens if you block diffusion of protons thru F0
concentration of protons builds up, levelling off of O2 consumption
why do we see leveling off of o2 consumption and atp synthesis
when u block ATP synthase’s ability to allow diffusion of protons thru F0, you prevent further flow of electrons. stuff builds up, more and more accumulation, electrons back up, no further electron transport along ETC, everything stops. leveling off of O2 consumption
describe how we showed ATP synthesis (ox phos) and ETC are linked
not enough just to have reactants for ATP or just electron, we need BOTH
what happens if you block diffusion of protons
also block electron transport
how do wee see this blocking of electron transport
leveling off of O2 consumption
what is an uncoupler of electron flow
separates/uncouples process of electron transport from ATP synthesis
example of uncoupler
dinitrophenol (DNP)
what happens if you block electron transfer
block ATP synthesis
what do we need to get respiration AND ATP synthesis
need both a source of electrons (succinate) and ADP + Pi
how do we know electron transport and ox phos are dependent?
experiments; isolated mitochondria from muscle. measure O2 consumption and ATP synthesis
what happens if you just add reactants for ATP synthesis (ADP and Pi)
nothing; tiny increase in O2 consumption, no increase in ATP synthesis
when do you get increase in both ATP synthesis and O2 consumption
when you add ADP & Pi AND succinate (source of electrons)
what do you see when you block electron transport
leveling off of ATP synthesis and O2 consumption
what blocks ATP synthase
venturicidin and oligomysin
describe how uncouplers work
venturicidin/oligomycin block ATP synthase, H builds up. takes too much energy to pump H across gradient. uncouplers dissipate the gradient (carry protons from P to N), allows ETC to begin again
can you have ETC without ATP synthesis
yup
what is DNP
hydrophobic, dissociable proton
what can DNP do
dissipate H+ gradient by carrying H+ across inner mitochondrial membrane
what happens if you add DNP
destroy proton gradient
what happens if you destroy gradient
ETC can continue
how do we see this uncoupling of DNP on the graph
increase in O2 consumption, but ATP synthesis is still same line (levels off)
when does uncoupling of ETC from ATP synthesis occur in real world
hibernation in mammals (some plants also)
do hibernating bears have DNP
nope
what do bears have
brown fat
what is brown fat
high in mitochondria [called brown fat cuz of cytochromes present in mitochondria causing it to look brown]
when does brown fat accummulate
winter approaches, temperature drops
what else do bears have
important protein called thermogenin
what does thermogenin do
uncouples ATP synthesis from ETC
why does this matter in hibernating animals?
aren’t expending as much energy, but they need heat.
what does natural uncoupling lead to
allows more E derived from flow of e- thru ETC, to be released as heat
what else is brown fat in
babies
what does brown fat serve as
significant heat source
what protein does uncoupling involve
thermogenin
what happens as winter approaches
hibernating mammals increase expression of thermogenin
what is thermogenin
proton pore protein
what does pore protein thermogenin do
allows protons to flow down gradient
how does thermogenin function
functions like F0, but doesn’t have F1 component (so not synthesizing ATP)
what does lack of F1 mean
it’s not synthesizing ATP
what does thermogenin essentially function as
a bypass proton pore
what does increasing thermogenin expression do
more proteins will diffuse thru thermogenin vs ATP synthase, simply b/c there’s more thermogenin around
what does thermogenin do
dissipate the gradient (like DNP)
how does thermogenin dissipate gradient
by its proton pore function
what happens as more protons diffuse thru thermogenin
more E from electron flow is generated in form of heat
