ATP Synthase Lecture 25 PDF

Electron Flow in the Respiratory Chain ATP synthesis  The Chemiosmotic Hypothesis Proposed by Peter Mitchell in the 1960’s (Nobel Prize 1978) Chemiosmotic = Chemical reaction + transport process Theory: electron transport and ATP synthesis are coupled by a proton...

Electron Flow in the Respiratory Chain ATP synthesis  The Chemiosmotic Hypothesis Proposed by Peter Mitchell in the 1960’s (Nobel Prize 1978) Chemiosmotic = Chemical reaction + transport process Theory: electron transport and ATP synthesis are coupled by a proton gradient across the inner mitochondrial membrane Glynn Research Mitchell’s postulates for chemiosmosis Institute, Bodmin, Cornwall Intact inner mitochondrial membrane is required Electron transport through the ETC generates a proton gradient ATP synthase catalyses phosphorylation of ADP - reaction driven by movement of H+ across the IMM into the matrix  ATP + H2O ADP + Pi ΔG = - 40 to -50 kJ/mol ADP + Pi ATP + H2O ΔG = + 40 to +50 kJ/mol How to reverse direction? Anisotropic distribution of H2O: Vectorially separate H+ from OH- so that reaction is pulled in “wrong” direction. Suggested protons must be “pumped” creating a gradient.  Chemiosmotic Model for ATP Synthesis Electron transport sets up a proton-motive force. Energy of proton-motive force drives synthesis of ATP.  ATPase – Complex V Efraim Racker, 1960’s F1 F0 Fo Oligomycin sensitive -antibiotic uncoupler Similar to bacterial and plant ATPase  ATP Synthase = Complex V (Fo F1 ATPase, or Proton- Stator: a b2 δ translocating ATP Synthase) ADP3- + Pi2- + H+ ATP4- + H2O F1 85 A complex with ATPase / ATP synthase activity. 8 subunits: 3. Rotor: c8-15 γ ε Fo = hydrophobic H+ channel embedded in IMM (subunits a and c). 1 H+ per C subunit, so stoichiometry of H+s needed per ATP varies between species  H+ PDB 1A91 Asp61 Fo proposed a ring of mechanism: subunit c subunits H+ Fo subunit c Rotation of ring of c subunits involves: concerted swiveling movements of the c- subunit helix (protonation/deprotonation of Asp61), a-subunit helices - Arg 210 transfers H+ to or from Asp61, protons are passed from or to each half- channel of subunit a. γ subunit is asymmetric puts force on each β subunit as it rotates. Subunit “a” has 2 half-channels Each H+ enters 1st half-channel of “a” Binds to “c” at Asp 61 H+ goes all the way around Released to 2nd half-channel back in “a” Evidence for Rotation Actin filament with fluorescent tag – microscopy t interval 133 ms  F1 catalyzes ADP + Pi ATP Hexamer arranged in three αβ dimers Each αβ dimer - three different conformations: – tight: catalyzes ATP loose: binding formation and binds ADP and Pi product open: empty The ATPase activity is in the  subunits. The 3  subunits = chemically the same but different conformations: O (open) Poor binding [to ADP + Pi or ATP] L (loose) binding [to ADP + Pi] T (Tight) binding [to ATP] The rotating  subunit (arrow) forces the conformational shift of: O  L  T  O ….. Hardest Step is not making ATP, but forcing the T conformation to release it, i.e. T → O The “binding-change” model for ATP synthase The “binding-change” model for ATP synthase Revolving  subunit “kicks” the  to make it give up the ATP  Coupling Proton Translocation to ATP Synthesis - Summary Proton translocation causes a rotation of the F0 subunit and the central shaft . This causes a conformational change within all the three  pairs. The conformational change in one of the three pairs promotes condensation of ADP and Pi into ATP. https://youtu.be/5DLHsyrrUOI Transport of ADP and Pi into the Matrix Translocation of 4th H+ per ATP - required to facilitate cotransport of substrates into, and products out of, the mitochondria.  ATP Synthesis Can Be Uncoupled from ETC to Generate Heat Uncoupling protein 1 (UCP-1) in babies (Brown Adipose Tissue, many mitochondria) Hibernating animals  Alternative oxidase (AOX) – CN- resistant respiration in plants Thermogenesis in the Arum spadix Why? Elevated temp. causes volatiles to evaporate - Smell of rotting flesh – attracts insect pollinators Cycad cones Corpse Flower Amorphophallus titanium