General Biochemistry 2: Lecture 9 (ATP Synthesis)

LECTURE 9: ATP SYNTHESIS

Energy coupling- free energy released by electron transport must be saved in a form that atp synthase can use.

chemiosmotic theory (1961, peter mitchell)- ETàcoupled to pumping of H⁺ from mitochondria matrix across inner membrane to outside à create H⁺ electrochemical potential gradient across inner membrane à this gradient is energy source which drives atp synthesis

-inner membrane permeability- requires intact inner membrane, impermeable to ions like H⁺, OH-, K+, Cl- (permeable to CO₂, O₂, H₂O)

-electrochemical gradient- ET = H⁺ transport from matrix to outside, creating big gradient across inner membrane

-Potential = 0.168 V (more negative inside);

-pH of matrix = 0.75 units higher than intermembrane space,

-ΔG(transport H⁺ outside) = 21.5kJ/mol

-2,4dinitrophenol (DNP)- compounds that increase permeability of inner membrane to H⁺ allow ET to proceed, but inhibit ATP synthesis. Thus, oxidation (or ET) and phosphorylation can be uncoupled by uncouplers like 2,4-dintrophenol (DNP)

p/o ratio- ratio of atp synthesized to oxygen reduced

p/2e ratio-ratio of atp synthesized to 2-e reduction of some e⁻ acceptor other than o

-FADH₂/O₂(no agent or amytal)=2, (w CN- or DNP)=0

-FADH₂/Fe(w CN-)-3 (no agent or CN-)=1

-NADH/O₂(w CN-)=0

-NADH/Fe(CN6)-3=2

-pH gradient- atp is made by mitochondria/chloroplast when pH gradient artifically generated without ET.

-bacteriorhodopsin-purple membrane protein from halobacteria; pumps H⁺ when illuminated; synthetic lipid vesicles w BR and beef heart mitochondrial ATPase can make ATP when illuminated.

-asymmetric orientation- NADH-Q reductase, Q-Cyt c reductase, Cyt c, cyt c oxidase, ATP synthase all asymmetrically oritented (used mitoc/submitoc particles w inner membrane inside out)

-Efraim Racker- discovered F0 and F1

1.F1- water soluble, peripheral membrane protein; atp hydrolysis in isolated soluble form

-a3b3yE, active in atp synthesis in synthase assembly

2.F0-water insoluble, transmembrane protein; forms proton channel.

-a1b2c9-12 in e. coli, and some addition subunits in mitochondrial F0

3. Binding Change Mechanism- shape change takes place when T has ATP and L has ADP,Pi

-L,O,T(atp)àL(adp,p),O,T(atp) + energyà L,O(ATP),T(adp,p) + ATP,waterà L,O,T(atp)

-shape change takes place when T site has ATP and L site has ADP,Pi; results in conversion of LàT, TàO, OàL;

-shape change requires energy, transmiited to catalytic α3β3 assembly via ye assembly.

-atp synthesize at new t site, original atp released from new o site