Chapter 14: The Electron Transport Chain

Electron Transport Chain takes place where?

in the mitochondria. the inner membrane creates two separate compartments in the mitochondria:

matrix--inner compartment that contains many enzymes and cofactors

intermembrane space--space between the two membranes.

the membrane with the matrix on the bottom and intermembrane space on the top contains 4 large protein complexes that transport electrons. each complex contains different proteins (i.e. complex 1 has 46 proteins).

Proteins that make up the 4 different complexes in the ETC

1. flavoproteins

2. cytochromes

3. Fe-S proteins

4. ubiquinones (coenzyme Q)

5. copper-containing proteins

flavoproteins

proteins that contain either FAD or another flavin-containing molecule called FMN (flavin mononucleotide). both of these are derived from riboflavin (vitamins!)

cytochromes

broad class of electron transfer proteins that contain an iron (Fe+3) ion within a heme group like hemoglobin. very abundant in the ETC (in every complex EXCEPT #1). the Fe ion accepts the electrons. most of these are contained within the large complexes; an exception to this is in cytochrome c: a small protein that moves between complexes outside of the membrane in the intermembrane space.

Fe-S proteins:

electron transfer proteins that also contain an iron ion but not within a heme group. the iron is instead bound to sulfur atoms; very abundant.

ubiquinones (coenzyme Q)

fat-soluble, electron transfer proteins that move freely between complexes in the membrane. can transfer 2 electrons at the same time.

copper-containing proteins:

similar to cytochromes and Fe-S proteins except they have Cu ions instead of Fe; the Cu accepts the e-

Steps of how electrons go through the ETC

1. e- are transferred from NADH to complex 1 (NADH dehydrogenase), oxidizing NADH to NAD+; the FMN in the complex are reduced. the e- are moved between many proteins in the complex, and in the process H+ ions are shuttled from the matrix to the intermembrane space

2. e- are transferred from complex 1 to coenzyme Q (ubiquinone). proteins in complex 1 are oxidized; ones in coQ are reduced during the transfer. coQ takes e- to Complex 3, oxidizing proteins in coQ and reducing proteins in Complex 3. more H+ ions move into intermembrane space as e- transfer to Complex 3.

3. e- are transferred from complex 2 (succinate dehydrogenase; part of the kreb's cycle that reduces FAD to FADH2) to coQ. these e- are then also transferred to complex 3 through coQ, moving more H+ ions in the process.

4. e- are transferred from complex 3 to mobile cytochrome C molecule (water soluble; not located in the membrane). cytoC takes the e- to complex 4, oxidizing proteins in cytoC and reducing them in complex 4. Cu-containing protein in complex 4 transfers the e- to O2 in the matrix, creating water. the movement of the e- moves even more H+ ions. O2 is the final e- acceptor; it is reduced.

How is ATP made in the ETC?

there are more H+ ions in the intermembrane space than the matrix due to the shuttling of so many ions throughout the ETC. These ions flow back into the matrix through ATP synthase w/ chemiosmosis. the enzyme uses the energy from the H+ flow to add a phosphate group to ADP to create ATP--called oxidative phosphorylation. this enzyme is a turbine that spins.

results in a production of 32-34 ATP. (only 4 produced in glycolysis and Kreb's). Boyer, Walker, and Skou won Nobel for this.

Mechanism of the movement of ATP Synthase

a proton binds to the rotor subunit, forming ionic bonds with neg. charged sidechains. this allows for the now neutral subunit to rotate into the hydrophobic lipid bilayer. proton exits out of channel after subunit completes 360 rotation. direction of rotation due to flow of proteins down the conc. gradient. energy stored in this gradient converted to mechanical rotational energy.

the central shaft rotates within the lollipop head (F1 ATPase). the head consists of three identical catalytic alpha-beta dimers. the shaft is asymmetric. as it rotates, it pushes the alpha-beta dimers through three conformations:

-binds ADP and iP

-catalyzes bond to make ATP

-releases ATP

we know this due to xray crystal structure that shows the dimers in three different conformations. the xray structures were static; single molecules too small to see in a light microscope.

Experimental Proof that ATP Synthase Rotates

a fluorescently-labeled long protein fiber (made of actin, part of the cytoskeleton) was attached to the tip of the shaft. you can see the fiber rotate in a light microscope while the enzyme works. ex. of single molecule biochemistry

How do we know the proton gradient across the membrane drives ATP synthesis?

purify ATP synthase and bateriorhodopsin, a bacterial protein that pumps protons across the membrane with energy from light. put both in lipid vesicle. shine light on it: the bacteriorhodopsin pumps the H+ into the vesicle and creates a gradient. ATP synthase only catalyzes ATP formation when light is shined. thus, the bacteriorhodopsin replaces the entire ET process. both create proton gradient; the proton gradient is enough to drive ATP synthesis.

Anaerobic Pathways for Carbohydrate Catabolism

many organisms can't use O2 as the final e- acceptor. must break down glucose w/o oxygen. these pathways generate much less ATP than aerobic pathways. two major pathways:

fermentation: only energy produced here is 2 ATP during glycolysis. convert pyruvate into lactic acid or alcohol, so glucose isn't broken down into water or carbon dioxide. use this process to make yogurt, beer, wine, etc.

anaerobic respiration: bacteria use this. nitrate, carbonate, sulfate are final e- acceptors. Krebs and ETC only partially works when oxygen is absent. more ATP produced here than in fermentation; still a very little.

Protein and Lipid Catabolism for Energy

fats can be broken down into fatty acids and glycerol components. glycerol is converted and inserted into glycolysis as G3P and fatty acids are converted to acetyl CoA, which goes into the Kreb's cycle.

proteins are broken down into amino acids which are then converted into molecules such as pyruvate.

ETC + Chemiosmosis in Bacteria and Plants

bacteria uses the same process in the ETC and chemiosmosis; they just use the plasma membrane instead of the mitochondrial membrane bc no mitochondria present.

plant chloroplasts use electrons from chlorophyll, final e- acceptor is NADP+. has different complexes but the same function.

Photosynthesis

produces glucose and oxygen from water and carbon dioxide. all O2 comes from plants. this process happens in the chloroplasts. stacks of thylakoid membranes within the chloroplast carry membrane protein complexes that carry this out. two parts: light reactions and light-independent reactions.

Light reactions of photosynthesis

takes place in the thylakoid membrane, with the thylakoid interior on top and stroma on the bottom. chlorophyll electrons in photosystem 2 are excited by light and reduces the carrier PQ. e- from water reduces chlorophyll again, resulting in O2.

PQ carries e- to cytochrome and plastocyanin carries them to photosystem 1. H+ is pumped in here. cytochrome pumps H+ exactly like complex 3 in mitochondrial e- transport. light re-excites e- in chlorophyll in PS 1, making them energetic enough to reduce ferredoxin and then NADP+ to make NADPH. ATP synthase then works like in mitochondria.

How do the photosystems of the chloroplasts work?

e- in the bonds of the ring around a magnesium ion are excited by visible light to more energetic orbitals. photosystems have an antenna with hundreds of chlorophylls and other small molecules that absorb light.

"resonance transfer" -- quantum mechanical effect that transfers energy from one molecule to another, eventually exciting e- in two special chlorophylls at the reaction center. these e- oxidize an e- acceptor.

an xray structure of PS 1 shows it's a trimer with proteins, chlorophylls, and carotenoids. close in on one monomer and find that reaction center is in the middle with Fe and S atoms at the center that carry e- given up by chlorophyll and pass them on.

the sole source of energy that enables the low entropy state in all of biology is the light captured by photosystems to excite e- so they can reduce other molecules.

The Calvin Cycle

light independent reactions of photosynthesis. uses ATP energy and e- from NADPH to reduce carbon dioxide and produce sugars.

uses carbon fixation: reduces atmospheric CO2 to produce other biological molecules. the enzyme RuBisCO does the carbon fixation. produces 18 ADP+Pi, one sugar, and 12 NADP+

Radioisotopic labeling of CO2 being converted to 3 phosphoglycerate

experiment by Melvin Calvin. used algae in a flask. pumped in 14_CO2 at time zero; 3 or 30 sec later, open stopcock to drain algae into boiling alcohol (stops enzymes). extract small molecules and spot on paper. separate small molecules by 2D paper chromatography: solvents wick through the paper and drag diff. molecules diff. distances. put x-ray film on paper and detect spots where 14_C molecules are.

tells us that the initial product of CO2 fixation is 3PG. later, the carbon from CO2 ends up in many molecules.

RuBisCO

a bad enzyme; catalyzes only 3 rxns/sec at max. need a lot of it; 50% of proteins in a leaf is this. active site only has 10-fold preference for binding CO2 than O2--often using O2 to oxidize the ribulose bisphosphate instead of CO2 which takes longer and more steps, consumes ATP, and releases CO2 (photorespiration).

thought to be remnant of evolution. photosynthesis appeared 3.5 billion years ago and no O2 in atmosphere at the time. 12 million years ago, new plants appeared with mechanisms to minimize photorespiration; corn, grasses, cacti, succulents--grow faster in hotter/drier environments due to this?