Mitochondrial electron transport chain Complex II succinate coenzyme Q reductase o aka flavoprotein 2 FP2 FAD covalently bound o four subunits including 2 Fe S proteins o one is succinate dehydrogenase from TCA cycle two TM CybL and CybS one more cytosolic Ip contains iron sulfur clusters o Three types of Fe S cluster 4Fe 4S 3Fe 4S 2Fe 2S Path 3 37 o Net reaction Other heme group fine tunes electronic properties succinate CoQ fumarate CoQH2 Doesn t pump H across membrane but shuttles e to CoQ for use by complex III Complex III mediates electron transport from coenzyme Q to cytochrome C o Aka UQ Cytochrome c Reductase o CoQH2 passes electrons to cyt c and pumps H in a unique redox cycle known as the Q cycle o The players The principal transmembrane protein in complex III is the b cytochrome with hemes bL and bH Cytochromes are one electron transfer agents CoQH2 is a lipid soluble electron carrier cyt c is a water soluble electron carrier The Q cycle 12 37 Q cycle phase 1 o QH2 binds complex III at cyt b subunit o QH2 reduces iron sulfur protein ISP 1 e forming Q releasing 2H ISP reduces Cyt c o Q reduces heme to form Q o Q diffuses to other side of membrane o gets re reduced to Q Q cycle phase 2 o another QH2 binds Goes through same pathway o reduces another Cyt c to form Q o Q reduces the other currently bound Q to form QH2 and Q Complex IV transfers electrons from cytochrome c to reduce oxygen on the matrix side o Cytochrome c Oxidase o overall rxn dimer with 13 different subunits per protomer just looking at reactants and products 4 H O2 4 cytochrome c Fe2 4 cytochrome c Fe3 2H2O if you also consider changes in H position 8 H matrix O2 4 cytochrome c Fe2 4 cytochrome c Fe3 2H2O 4 H intermembrane space note one cycle requires oxidation of 2 NADH Electrons from cyt c are used in a four electron reduction of O2 to produce 2 H2O Complex IV o Oxygen is thus terminal acceptor of electrons in the electron transport pathway o complex IV Cytochrome c oxidase utilizes 2 hemes a and a3 and 2 copper sites o mechanism involves changes in oxidation state of heme Fe and Cu also involves formation of tyrosyl radical O O bond split and one O bound to Cu II and one to Fe IV then cleaved off as H2O by addition of e and H o 2 H are transported across the inner mitochondrial membrane per NADH called vectorial protons Note for complex IV and complex I H are consumed in the matrix scalar protons in addition to being pumped out H consumed in matrix contribute to negative membrane potential that pull H into matrix during ATP production Overall e transport o Overall 10 protons pumped out per oxidation of 1 NADH o 4 H in complex I 4H in complex III and 2H in complex IV o Proton Gradient The chemiosmotic hypothesis Proton gradient across the inner membrane drives ATP synthesis Proposed by Peter Mitchell in 1961 Initially ridiculed but the won him a Nobel prize in 1978 Chemiosmotic hypothesis o Observations that led to the chemiosmotic hypothesis e transport generates H gradient inner mito membrane is impermeable to ions maintains eletrochemical gradient ATP synthesis requires intact inner membrane compounds that increase permeability of inner mito membrane inhibit ATP synthesis but do not stop electron transport o e transport and ATP synthesis become uncoupled proton motive force o Proton motive force energy for moving a proton from matrix to inter membrane space o 5 38 o If matrix pH 8 intermembrane pH 7 0 168V T 37 C o G 22 1 kJ mol matrix intermembrane space o so going opposite direction is 22 1 kJ mol o 50 kJ mol for synthesizing ATP from ADP so need at least 2 H biochemical measurements indicate need 3 H per 1 ATP synthesis How does a proton gradient drive the synthesis of ATP o Proton transport through the ATP synthase drives ATP synthesis o ATP synthase aka F1F0 ATPase consists of two parts F1 and F0 latter was originally named Fo for its inhibition by oligomycin F1F0 ATPasae structure o F1 consists of five polypeptides and F1 is catalytic subunit are nearly identical arranged in pseudo threefold symmetry both and can bind ADP ATP but only does catalysis together dimers adopt three different conformations defined by their affinity for ADP ATP o low O state medium L state and high affinity T state forms rod going through the center of and wrap around the base of rotational catalysis o the three subunits of F1 are driven between different conformations by rotations of F0 that are transduced through three steps of ATP synthesis turn start at loose L conformation bind ADP rotation drives conformation to tight T conformation forms ATP further rotation drives conformation to open O conformation releases ATP o rotation of rotor driven by transport of H proton flow o protons carried across membrane through combined action of a subunit and c subunit H enter through pore in a subunit H neutralize a neg charged Asp in middle of TM helix of c subunit allows rotor to otherwise would have unfavorable charge in the middle of the membrane apparently entry point and exit point of H on the a subunit are dislocated from each other o protons only released on matrix side after full turn of rotor ATP ADP translocase mediates the movement of ATP ADP o ATP out ADP in through a translocase o ATP movement out is favored because the cytosol is relative to the matrix o But ATP out and ADP and Pi in is net movement of a negative charge in equivalent to a H going in to be electrostatically neutral exchange o So every ATP transported out costs one H o One ATP synthesis costs about 3 H o Thus making and exporting 1 ATP 4H How many ATP can be made per electron pair in e transport o The e transport chain yields 10 H pumped out per electron pair from NADH to oxygen o 4 H flow back into matrix per ATP to cytosol 10 4 2 5 for electrons entering as NADH also called P O ratio for number of ATP produced per oxygen reduced o For electrons entering as succinate FADH2 about 6 H pumped per electron pair to 6 4 1 5 for electrons entering as succinate Utilizing NADH from glycolysis o amount of ATP produced depends on the shuttle used to move NADH equivalents into o Malate aspartate shuttle uses malate to carry electrons across the membrane yields 2 5 ATP per NADH o Glycerophosphate shuttle stores electrons in glycerol 3 P which transfers electrons to oxygen matrix FAD yields 1 5 ATP per cytosolic NADH o so 32 ATP per glucose if malate Asp shuttle used 2 5 ATP NADH 10 NADH glucose 1 5 ATP FADH2 2 FADH2 glucose 2 ATP glucose TCA 2ATP glucose