Biochem Exam 3 Ch. 20 (22-.4 will be separate)

a mobile electron carrier

Coenzyme Q (called Ubiquinone, UQ or Co Q) is what? There are 3 oxidation states of this as shown, be able to draw them, especially quinone and *quinol(specifically)*.

flow back into the matrix

Coenzyme Q collects electrons from NADH-UQ reductase and succinate-UQ reductase and delivers them (by diffusion through the membrane core) to UQ-cyt c reductase. Cytochrome c is water soluble and moves freely in the intermembrane space, carrying electrons from UQ-cyt c reductase to cytochrome c oxidase. In the process of these electron transfers, protons are driven across the inner membrane (from the matrix side to the intermembrane space). The proton gradient generated by electron transport represents an enormous source of potential energy. This potential energy is used to synthesize ATP as protons do what?

view picture

Complex 1 and 2 as well as flavoproteins contributes to ubiquinal(UQH2) generation in pool

FMN, NADH-UQ reductase is a flavoprotein.

Complex 1 depends on what? (yes, this horse is getting beaten)

by partitioning some of the apoptotic activator molecules, e.g., cytochrome c Oxidation of bound cardiolipins releases cytochrome c from the inner membrane Opening of pores in the outer membrane releases cytochrome c from the mitochondria Binding of cytochrome c to Apaf-1 in the cytosol leads to assembly of apoptosomes, thus triggering the events of apoptosis (Maybe TMI but know the fist one)

How Do Mitochondria Mediate Apoptosis?

Proton diffusion through the ATP synthase

How Does a Proton Gradient Drive the Synthesis of ATP?

1 proton 3 protons 4 protons

So every ATP transported out costs _______ One ATP synthesis costs about _________ Thus, making and exporting 1 ATP = ________ (protons)

cardiolipin

Substantial evidence points to a requirement for _______________ in the assembly and stability of mitochondrial supercomplexes. For example, association of Complexes III and IV requires the presence of "this". Studies by Carola Hunte and colleagues show that "this" stabilizes supercomplexes by neutralizing the charges of lysine residues at the binding interface between Complex III and Complex IV.

32.0 34.2

______ATP per glucose if glycerol-3-P shuttle used ______ATP per glucose if malate-Asp shuttle used

Oxygen

__________ is the terminal acceptor of electrons in the electron transport pathway

the electron-transferring flavoprotein, which transfers electrons from the flavoprotein-linked step of fatty acyl-CoA dehydrogenase, and sn-glycerophosphate dehydrogenase.

there are two other ways to feed electrons to UQ what are they?

you are breaking or forming covalent bonds

Arrhenius equation only applies when what?

Complex I Complex IV ATP synthase

Rotenone , Amytal, and Demerol inhibits _____________ Cyanide, azide and CO inhibit _____________, binding tightly to the ferric form (Fe3+) of a3 Oligomycin inhibits __________

NADH to the enzyme on the matrix side of the inner mitochondrial membrane and transfer of electrons from NADH to tightly bound FMN. (remember O.i.L.R.i.G.)

(Complex 1) Although the precise mechanism of the NADH-UQ reductase is unknown, the first step involves binding of what?

the reduced [FMNH2] to a series of Fe-S proteins (including both 2Fe-2S and 4Fe-4S clusters)

(Complex 1) The second step involves the transfer of electrons from what to what?

two electrons from iron-sulfur clusters to coenzyme Q. Coenzyme Q is a mobile electron carrier. Its isoprenoid tail makes it highly hydrophobic, and it diffuses freely in the hydrophobic core of the inner mitochondrial membrane. As a result, it shuttles electrons from Complexes I and II to Complex III.

(Complex 1) The final step of the reaction involves the transfer of what to what? describe how the mobile electron carrier is able to be in this case.

FAD

(Complex II) The 2 flavoproteins (Fp) contain an _______ covalently bound to a His residue

4Fe-4S, 3Fe-4S, 2Fe-2S succinate → FADH2 → 2Fe2+ → UQH2

(Complex II) What are the three types of Iron sulfur clusters in Ip of Complex II? what is succinate's path?

Fe-S centers UQ (Co Q)

(Complex II) When succinate is converted to fumarate in the TCA cycle, concomitant reduction of bound [FAD] to [FADH2] occurs in succinate dehydrogenase. This [FADH2] transfers its electrons immediately to __________, which pass them on to ______.

a molecule of UQH2 from this pool diffuses to a site (called Qp) on Complex III near the cytosolic face of the membrane.

(Complex III) A large pool of UQ and UQH2 exists in the inner mitochondrial membrane. The Q cycle is initiated when what happens?

proton transport across the inner mitochondrial membrane.

(Complex III) As with Complex I, passage of electrons through the Q cycle of Complex III is accompanied by

Rieske cytochrome c1 2 UQ- to UQ

(Complex III) Oxidation of this UQH2 occurs in two steps. First, an electron from UQH2 is transferred to the __________ protein and then to _________________. This releases ______ H+ to the cytosol and leaves UQ- , a semiquinone anion form of UQ, at the Qp site. The second electron is then transferred to the bL heme, converting ______ to ______.

b cytochrome The b cytochromes contain iron protoporphyrin IX , the same heme found in hemoglobin and myoglobin. (Cytochromes, like Fe in Fe-S clusters, are one- electron transfer agents )

(Complex III) The principal transmembrane protein in complex III is the ____________ - with hemes bL and bH (H and L here refer to high and low reduction potential.) what is significant about this transmembrane protein?

cytochrome bc1 complex

(Complex III) The structure of the UQ-cyt c reductase, also known as the _______________________.

hemes (a and a3) Copper sites (2 CuA center and 1 CuB site)

(Complex IV) Cytochrome c oxidase utilizes 2 ________ and 2 _________

4 Cytochrome C

(Complex IV) Electron transfer through Complex IV begins with binding of cytochrome c to the b-barrel of subunit II. ________ electrons are transferred sequentially (one each from four molecules of ________________) first to the CuA center, next to heme a, and finally to the CuB/heme a3 active site, where O2 is reduced to H2O

4 4

(Complex IV) How many protons participate in O2 reduction? How many protons are transported in each catalytic cycle?

this is the catalytic site where O2 binds and is reduced.

(Complex IV) The Fe atom in heme a3 is five coordinate, with four ligands from the heme plane and one from His376. This leaves a sixth position free, what is sOoOoo special about this site?

spontaneous (net -) "driving force for biological systems" non-spontaneous (net +) reorganization energy = inner sphere (amino acids and other parts of enzymes; this is where issues arise) + outer sphere {lithium ion doesnt have issues with this, however the ETC does} Battery: Fe³+ →Cd¹+ (get e- from point A to B) Biological system: bc1's Fe²+ ← Cyt c Fe (Needs to get e- from point A to B but need to minimize loss of e- efficiency, survival ) loss (entropy) produces ROS → cytochrome c → apoptosis

(redox potential E⁰) ∆G= -nFE⁰ or.... ∆G = -(#mole-)(96485 c/mole-) if E⁰ is + then the reaction is what? if - then what? Describe reorganization energy.

NADH FADH2 33.6

10 H+/NADH and 10 H+/3 ATP = 3 ATP/_________ 6 H+/succ and 10 H+/3ATP = ~ 1.8ATP/__________ (10 NADH + 2 [FADH2])/glucose → ________ ATP

Che-Che-CHECK IT OUUUUT!

A model for the electron-transport pathway in the mitochondrial inner membrane. UQ/UQH2 and cyt c are mobile carriers and transfer electrons between the complexes

oxidative phosphorylation (The proton gradient established across the inner mitochondrial membrane drives the synthesis of ATP) O2 (molecular oxygen)

ATP made in glycolysis and the TCA cycle is the result of substrate-level phosphorylation, NADH-dependent ATP synthesis is the result of _________________________. Electrons stored in the form of the reduced coenzymes, NADH or [FADH2], are passed through an elaborate and highly organized chain of proteins and coenzymes, the so-called electron-transport chain, finally reaching _________________, the terminal electron acceptor.

+ - (But ATP out and ADP in is net movement of a negative charge out - equivalent to a H+ going in)

ATP movement out is favored because the cytosol is "_____" relative to the "______" matrix

WD40

Binding of cytochrome c to the __________ domains and ATP hydrolysis unlocks Apaf-1 to form the semi-open conformation. Nucleotide exchange leads to oligomerization and apoptosome formation.

proton channels and the redox centers. (Subunits I, II, and III are common to most organisms. This minimal complex is sufficient to carry out both oxygen reduction and proton transport more specifically)

Bovine cytochrome c oxidase consists of 13 subunits. The 3 largest subunits - I (purple), II (yellow), and III (blue) - contain the what?

zero

Boyer's experiments showed that ¹⁸O could be incorporated into all four positions of phosphate, demonstrating that the free energy change for ATP formation from enzyme-bound ADP + Pi is close to what?

NADH Coenzyme Q (also called ubiquinone or UQ or CoQ) NADH dehydrogenase. 4, 2 NADH (donates 2e- and 2H+ to..) → FMN (reduced to FMNH2 needs 2H+ to bind the 2e- to it ) → Fe-S → UQ (also uses 2H+ to pick up those same 2e- converting to qubiquinol UQH2) (FMN is a flavin mononucleotide capable of 2e- pick up) (Complex I carries out electron transfer from NADH to CoEQ...Reductase and dehydrogenase) it is a proton pump (Complex I is 45 subunits and 980 kD)

Complex I Oxidizes _________ and Reduces ______________. Another name for this complex is what? _______ protons transported out per ______ electrons what is the electron path?

matrix cytosol

Complex I Transports Protons From the _________ to the ___________

Transports Protons from the Matrix to the Cytosol. The oxidation of one NADH and the reduction of one UQ by NADH-UQ reductase results in the net transport of protons from the matrix side to the cytosolic side of the inner membrane.

Complex I functional overview

Succinate (into fumarate) Coenzyme Q (ubiquinone, UQ or Co Q) which is given the 2e- through fe-s clusters as well as FAD being reduced to FADH2 Succinate→fumarate FAD→FADH2→FAD (bc it is kicking off 2H+ into the wild and 2e-, that travel through the Fe-S clusters to qubininone where it is reduced to ubiquinol (QH2) which shuttles on to my wayward son (complex III) (This complex does not pump protons and is involved in TCA and is called succinate reductase)

Complex II Oxidizes ____________ and Reduces ______________.

succinate dehydrogenase Complex II Oxidizes Succinate and Reduces Coenzyme Q (UQH2) succinate + UQ → fumarate + UQH2

Complex II is perhaps better known by its other name—______________________, the only TCA cycle enzyme that is an integral membrane protein in the inner mitochondrial membrane. What is the net reaction?

Electron Transport from Coenzyme Q to Cytochrome c reduced coenzyme Q (UQH2) passes its electrons to cytochrome c via a unique redox pathway known as the Q cycle.

Complex III Mediates what? how does this happen generally speaking?

UQH2 cytochrome c (note, this is different from cytochrome c₁ who carries 1 heme group) Complex IV (cytochrome c oxidase) molecular oxygen (purpose of this is to transfer 2 high energy electrons from ubiquinol to another carrier molecule called cytochrome c)

Complex III oxidizes __________ while reducing the water soluble ______________, which in turn is the substrate for ______________, that is also responsible for reducing what?

Cytochrome c Reduce Oxygen on the Matrix Side Cytochrome c Oxidase because it accepts electrons from cytochrome c and directs them to the four-electron reduction of O2 to form H2O (imagine that...)

Complex IV Transfers Electrons from _______________ to ______________. This complex is also called what? Why?

inner mitochondrial membrane

Complex IV transports H+ across the what?

cytochrome c (a mobile electron carrier)

Electrons traversing Complex III are passed through cytochrome c1 to _________________ because it is water soluble.

respirasomes

For many years, the complexes of the electron transport chain were thought to exist and function independently in the mitochondrial inner membrane However, growing experimental evidence supports the existence of multimeric supercomplexes of the four electron transport complexes Also known as _____________, these complexes may represent functional states

electrochemical

In Mitchell's chemiosmotic hypothesis, protons are driven across the membrane from the matrix to the intermembrane space and cytosol by the events of electron transport. This mechanism stores the energy of electron transport in a(an) _______________ potential.

2 protons UQH2 (two UQH2 are taken from the UQ/UQH2 pool and one is returned; the Q cycle accomplishes the net oxidation of one UQH2.)

In this latter half of the Q cycle, however, the bH electron is transferred to the semiquinone anion, UQ- , at the Qn site. With the addition of ______________ from the mitochondrial matrix, this produces a molecule of __________, which is released from the Qn site and returns to the coenzyme Q pool, completing the Q cycle.

cytosolic doesn't cross

Most NADH used in electron transport is ___________ and NADH ___________ the inner mitochondrial membrane

III

Mutation studies show that Complex ______ is required to maintain Complex I in the mitochondria of human and mouse cells in culture, and Complex IV is necessary for proper assembly of Complex I in mouse fibroblasts.

three

Paul Boyer proposed that, at any instant: the three β-subunits of F1 exist in ________ different conformations these different states represent the three steps of ATP synthesis each site steps through these conformations or states to make ATP In Boyer's binding change mechanism, the three catalytic sites thus cycle through the three intermediate states of ATP synthesis

Mitchell's chemiosmotic hypothesis.

Peter Mitchell, a British biochemist, proposed that the energy stored in a proton gradient across the inner mitochondrial membrane by electron transport drives the synthesis of ATP in cells. The proposal became known as what?

F0 Motor

Proton Flow Through _____ Drives Rotation of the _________ and Synthesis of ATP

Low, High, Active

Protons are being moved from a _______ concentration to a ________ concentration using _______ transport.

inlet c-ring rotor β

Protons entering the _________ half-channel in the α-subunit are transferred to binding sites on c-subunits. Rotation of the ____________ delivers protons to the outlet half-channel in the α-subunit. Flow of protons through the structure turns the ______ and drives the cycle of conformational changes in ____ that synthesize ATP.

sulfur atoms coordinates the iron.

Related to the structure of mitochondrial cytochrome c. The heme is at the center of the structure. It is covalently linked to the protein via two ___________. A third sulfur from a methionine residue does what?

F0 F1

The ATP synthase consists of two parts: F1 and F0 ____ forms the transmembrane pore or channel through which protons move to drive ATP synthesis Flow of protons from the a-subunit through the c-subunits turns the rotor and drives the cycle of conformation changes in the α- and β-subunits of ______ that synthesize ATP.

3

The Catalytic Sites of ATP Synthase Adopt _________ Different Conformations

NADH mitochondrial reduction of [FAD].

The Glycerophosphate Shuttle Ensures Efficient Use of Cytosolic _________ The glycerophosphate shuttle couples cytosolic oxidation of NADH with what?

open (O) loose (L) tight (T)

The binding change mechanism for ATP synthesis by ATP synthase. This model assumes that F1 has three interacting and conformationally distinct active sites: an ________________ conformation with almost no affinity for ligands, a ____________ conformation with low affinity for ligands, and a _____________ conformation with high affinity for ligands.

P (for positive) N (for negative)

The cytosolic side, where H+ accumulates, is referred to as the _______ face; similarly, the matrix side is the _________ face.

10 protons 3 ATP 3.7 Protons NADH 6 Protons succinate

The e- transport chain yields _____ H+ pumped out per electron pair from NADH to oxygen 8 H+ per turn of c₈ F₀ rotor → ____ ATP _____ H+ flow back into matrix per ATP to cytosol 10/3.7 = 2.7 ATP for electrons entering as _______ For electrons entering as succinate (FADH2), about ____ H+ pumped per electron pair to oxygen 6/3.7 = 1.6 ATP for electrons entering as ____________

bH UQ UQ-

The electron on the bL heme facing the cytosolic side of the membrane is now passed to the ____ heme on the matrix side of the membrane. The electron is then passed from this to a molecule of _____ at a second quinone-binding site, Qn, converting this to ______ . The resulting product remains firmly bound to the Qn site. This completes the first half of the Q cycle

cytosolic Electrons Copper and heme

The electron-transfer pathway for cytochrome c oxidase. Cytochrome c binds on the ___________ face, transferring ___________ through the __________ and __________ centers to reduce O2 on the matrix side of the membrane.

the oxidized, semiquinone, and reduced states NADH and the Fe-S proteins.

The flavin of FMN has three redox states—what are they? It can act as either a one-electron or a two-electron transfer agent and may serve as a critical link between what?

have an idea

The free energy difference for protons across the inner mitochondrial membrane includes a term for the concentration difference and a term for the electrical potential: c1 and c2 are proton concentrations on the two sides of the membrane, Z is the proton charge, F is Faraday's constant, and Δψ is the potential difference across the membrane

review this

The opening of pores in the outer membrane, induced by a variety of triggering agents, releases cytochrome c to the cytosol, where it initiates the events of apoptosis. Apaf-1 is a multidomain protein, consisting of an N-terminal CARD, a nucleotide-binding and oligomerization domain (NOD), and several WD40 domains Binding of cytochrome c to the WD40 domains and ATP hydrolysis unlocks Apaf-1 to form the semi-open conformation. Nucleotide exchange leads to oligomerization and apoptosome formation. A model of the apoptosome, a wheel-like structure with molecules of cytochrome c bound to the WD40 domains, which extend outward like spokes.

UQH2

The second half of the cycle is similar to the first half, with a second molecule of __________ oxidized at the Qp site, one electron being passed to cytochrome c1 and the other transferred to heme bL and then to heme bH.

Matrix

The space inside the inner mitochondrial membrane is called the _______, and it contains most of the enzymes of the TCA cycle and fatty acid oxidation.

1) A lipid-soluble coenzyme (UQ, CoQ) and a water-soluble protein (cytochrome c) shuttle between protein complexes 2) I and II to complex IV

There are 4 protein complexes in the inner mitochondrial membrane: 1) What shuttles between them? 2) Electrons generally fall in energy through the chain - from what to what complexes?

lipid water

UQH2 is a _______ -soluble electron carrier Cytochrome c is a ________ -soluble electron carrier

proton

Uncouplers are hydrophobic molecules with a dissociable __________ They shuttle back and forth across the membrane, carrying protons to dissipate the gradient

dissipating the proton gradient

Uncouplers disrupt the tight coupling between electron transport and oxidative phosphorylation by what?

UQ-cytochrome c reductase (UQ-cyt c reductase), (involving three different cytochromes and an Fe-S protein.)

What is complex III also called?

in or at the inner mitochondrial membrane (The typical mitochondrion is about 0.5 + or - 0.3 micron in diameter and from 0.5 micron to several microns long; its overall shape is sensitive to metabolic conditions in the cell. mammalian cells contain 800 to 2500 mitochondria)

Where in the Cell Do Electron Transport and Oxidative Phosphorylation Occur?

Complex I (contributes to Q pool) Complex II (contributes to Q pool) reduced coenzyme Q (UQH2) coenzyme Q-cytochrome c reductase (Complex III).

_____________ accepts electrons from NADH, serving as a link between glycolysis, the TCA cycle, fatty acid oxidation, and the electrontransport chain. _______________ includes succinate dehydrogenase and thus forms a direct link between the TCA cycle and electron transport. Both of these produce a common product, ______________________, which is the substrate for what?

ATP-ADP Translocase out in

_______________ Mediates the Movement of ATP & ADP Across the Mitochondrial Membrane ATP must be transported ____ of the mitochondria ADP _____ - through (what the very fist blank is)

Glycerophosphate Malate-aspartate

__________________ shuttle stores electrons in glycerol-3-P, which transfers electrons to FAD _______________ shuttle uses malate to carry electrons across the membrane (reversible)

(I) NADH-coenzyme Q reductase, (II) succinate-coenzyme Q reductase, (III) coenzyme Q-cytochrome c reductase, (IV) cytochrome c oxidase

what are the 4 parts of the Electron Transport Chain?