|Lecture Notes: 19 April||
With these electron transport components we can create a path for the transfer of electrons from substrate to oxygen, as above, but we want more than a wire: want to use this flow of electrons to run a couple of "motors" to pump protons across the inner mitohondrial membrane, forming an electrochemical gradient, as shown in Figure 14.6. [overhead, lower] Note that on this diagram the heights of the boxes indicates the change in standard potential or energy. From this we can see that complexes I and IV obviously have enough energy change to support the phosphorylation of ADP to ATP, while complex III is marginal and complex II obviously does not have sufficient energy change.
But how is this energy captured? Mitochondria appear to be using the energy of moving the electrons through this potential gradient to pump protons across the inner mitochondrial membrane. (Figure 14.1) [overhead] The resulting proton motive gradient can then be used to make ATP as we shall see later. Lets look at the four complexes and what seems to be occurring in each.
[Figure 14.7] Note that the electrons are received as a pair (a hydride ion) on NADH, but most carriers can only handle single electrons so FMN acts as a transducer, picking up a pair of electrons, but passing them on singly to the FeS centers in this complex. The electrons are then passed singly on to CoQ, which, like FMN can carry either pairs or single electrons. Four H+ are pumped across the inner mitochondrial membrane by complex I. (Note that formally the protons from NADH and H+ can be considered as going to FMNH2 and then QH2.)
[Figure 14.9] An electron pair enters this complex via FAD going to FADH2. Of course FAD has the same active portion as FMN so it can again carry electrons single or in pairs. FADH2 then passes its electrons on singly to FeS centers, which then pass them on to CoQ. No protons are pumped by this complex. Thus we see the difference in the number of ATPs which are produced by NADH vs. FADH2.
As we've seen before CoQ is a quinone with a long hydrocarbon tail. It exists as a pool of CoQ molecules dissolved in and diffusing through the lipid bilayer of the inner membrane. Both the quinone (oxidized) and quinol (reduced) form of the cofactor can "flip" in the membrane, thus the quinone ring can freely cross. CoQ can pick up electrons one at a time from either Complex I or II, then diffuse through the bilayer until collision with complex III allows it to pass them singly to that complex.
[Figure 14.11] This complex picks up single electrons from the CoQ pool, taking the quinol form in two steps to the quinone form of the coenzyme. One electron is passed on through FeS centers to Cytochrome c, while a second is recycled via cytochrome b (b566 & b560) to reduce Q to Q- in the Q-cycle (Figure 14.11) [overhead] . A second QH2 repeats the cycle and provides the second electron for the first recycled Q. The net result is the oxidation of one QH2 and the transport of four H+ for each pair of electrons flowing through the complex.
This cytochrome is a mobile protein carrier attached to the outer side of the inner mitochondrial membrane (a peripheral membrane protein). It transports single electrons from complex III to complex IV.
[Figure 14.13] This complex picks up electrons singly from cytochrome c and transfers them via cytochrome a-CuA to cytochrome a3-CuB where they are passed on to oxygen. Note that four electrons are needed for each oxygen molecule, and that the equivalent of two H+ are transferred out of the matrix for each pair of electrons (4 protons/oxygen). The high G for this complex, which would allow a significantly larger number of protons to be pumped, assures the completion of the overall reaction of oxygen and NADH to give water.
(For an insider's review and evidence on ETS and OxPhos see M. Saraste Science 283 (5 March 1999) pp 1488-93)
(Complex V; F0F1 ATP synthase, Figure 14.14) [overhead]: ATP synthase uses the proton gradient to make ATP from ADP and Pi. It is bound to the inner membrane and has a characteristic knob and stalk structure as seen in electron micrographs. It can be broken into two multi protein components: The F1 component (the "knob") hydrolyses ATP when it is isolated by itself and is referred to as F1 ATPase. The F0 component is a membrane spanning proton channel. When the two components are linked the passage of protons through the channel is coupled to ATP synthesis. According to the binding-change mechanism there are three sites in the alpha3beta3 oligomer of the knob. At any given time the three sites are in three different conformations, as shown in Figure 14.15: open, loose, or tight. [overhead] Each site passes sequentially through the three conformations, apparently while physically rotating 120° for each change. Following one site: 1) ADP and Pi bind to the site in the open conformation. 2) Passage of 3 protons through the channel causes the alpha-beta oligomer to rotate 120° and change to the loose or L conformation, , holding the ADP and Pi (all three active sites to go to the next conformation simultaneously). 3) Passage of another 3 protons through the channel causes the alpha-beta oligomer to rotate 120° and change to the tight conformation with consequent condensation of ADP and Pi to ATP. 4) Passage of another 3 protons through the channel causes the alpha-beta oligomer to rotate 120° and change to the open conformation, releasing ATP. Note the net result of 3 protons/ATP.
(For an insider's review and evidence on ETS and OxPhos see M. Saraste Science 283 (5 March 1999) pp 1488-93; for ATPase 'motor' see Paul D. Boyer (18 Nov 1999) "What matkes ATP synthase spin?" Nature 402: 247-8.)
Last modified 9 May 2010