The Three Models:

Historical: 1975 marked the transition from the chemical coupling model to chemiosmotic model. In this particular case, reductionism and Okam's razor failed to work well; it actually impeded progress. Outlined below are the three models under consideration at the time:

1. The chemo-mechanical (or conformational coupling) model involved the passage of electrons that caused a conformation change in th e proteins involved. This energy was then used to facilitate the formation of ATP.

2. The chemical coupling model proposed a mechanism similar to the substrate-level phosphorylation seen in glycolysis. A phosphate group would combined with a reduced substituent, which when oxidized would create a phosphate group in a higher energy configuration.

3. The chemiosmotic model, which is now the accepted model, was first proposed in 1961. In it, Peter Mitchell proposed that:

- electron transport results in the pumping of protons out of the mitochondrion
- a proton gradient builds up on the outside of the mitochondrion
- the protons return via a channel in an ATP synthase to form ATP

Evidence for chemiosmotic model

A. pH outside of the mitochondrion is 1.4 units lower than inside (25 x)

B. ATP is synthesized when pH gradient is imposed on mitochondrion.

C. intact mitochondria are necessary for ATP synthesis (fragments won't work)

D. substances (like dinitrophenol, DNP) that carry protons across the inner membrane of mitochondria dissipate the proton gradient, and no ATP is synthesized.

Mitochondrial ATP synthase:

ATP synthase is a protein complex located in the inner mitochondrial membrane, which converts the energy in the proton gradient into energy stored in the form of phosphate anhydride bonds in ATP.

Certain developments in the study of oxidative phosphorylation produced some unusual nomenclature. In the efforts to purify the ATP synthase (in earlier time called the ATPase), mitochondrial fragments were sheared, producing two fractions. The original one, containing a water-soluble portion of the complex was called F1. It contains 5 different polypeptide chains identified as , , , , . At a later time the other protein portion of the synthase was isolated from the membrane fraction and called Fo. It contained the proton channel of the complex and consisted of 4 polypeptide chains.

ATP is formed by the ATP synthase even in the absence of a H⁺ gradient. The proton gradient functions only to release the ATP. The detailed mechanism of this process is not known, since X-ray structures have not been obtained.

Chemiosmotic thermodynamics:

Thermodynamic arguments were used most persuasively, prior to 1975 ,to prevent the acceptance of the chemiosmotic model. An examination of oxidative phosphorylation from this perspective will permit us to understand how the current model is supported.

Remember that pH outside the inner mitochondrial membrane is 1.4 units lower than inside the mitochondrion, the difference yields two types of potential:

electrical potential --> _m
concentration gradient --> pH

The difference in potential between inside and outside:

p = _m - ( 2.3 RT/F) pH = 0.14 volts - (0.059)(-1.4) = 0.224 V

This difference in voltage corresponds to 21 kJ/mole of protons. On the basis of two (may be as high as 12, the stoichiometry is still in question) protons pumped at each site in the electron transport chain the amount of energy is

2 moles of protons x 21 kJ/mole of protons = 41 kJ

G^o' for ATP hydrolysis is -30.5 kJ/mol, therefore the energy from two protons returning is more than adequate for synthesis of ATP.

Summary of energy production from glucose

glucose-Pi (formation)					-1

fructose 1,6-bisphosphate				-1

1,3-bisphosphoglycerate					+2

phosphoenolpyruvate					+2

NADH from glycolysis (2 x 3 ATP)		      +6

pyruvate --> Ac-CoA  (inside mito  2 x 3 ATP)		+6

TCA cycle   2 GTP/Ac					+2

6 NADH  (6 x 3 ATP)					+18

2 FADH2  (2 x 2 ATP) 			+4
Total	 +38

This corresponds to 42% efficiency, which is better than almost all man-made machines.

NADH + 1/2 O₂ + H+ <==> H₂O + NAD⁺ G^o' = - 220 kJ/mole
ADP + P_i + H+ <==> ATP + H₂O G^o' = + 30.5 kJ/mole

30.5 x 3 = 91.5/220 = 42%

Shuttle systems:

Since the mitochondrial inner membrane must be impervious to protons (H+), it is not surprising that other, larger charged molecules can't move across this membrane without specific transport proteins. These proteins function to move:

phosphate (in), ADP (in), and ATP (out)
malate in, phosphate out (a dicarboxylate shuttle system)
citrate in, malate out (a tricarboxylate shuttle system)
aspartate (in), glutamate (out)
a-KG (in), malate (out)

Questions

Answer choices:

a. 0

b. 1

c. 2

2 Ascorbate provides a bypass to the antimycin A inhibition of complex III, how many moles of ATP are formed per atom of oxygen converted to water?

Answer choices:

a. 0

b. 1

3 If complexes I, III, and IV were able to pump protons out of the mitochondrion in proportion the amount of energy generated in the redox reaction taking place at each site, which site would pump the largest number of protons?

Answer choices:

a. complex I

b. complex III

c. complex IV

4 If an Fe₂S₂ iron sulfur protein were denatured, which of the following chemical changes would most likely occur?

Answer choices:

a. hydrogen sulfide would form

b. cysteine would be released from the protein

c. the iron would remain in the ferrous oxidation state