1. Rotational Catalysis Mechanism Binding-change model of  subunit  Proton passage through F o  conformational change in  subunits  Rotation of the cylinder of c &  subunits  Every  rotation of 120 o   contacts with a different  subunit  induce conformational change   -empty conformation  Neighbors of  -empty  -ADP or  - ATP  One complete rotation of  subunits ; Cycle of  -ADP   - ATP   -empty  3 ATP synthesis  Opposite rotation of  subunit in a single F 1  ATP hydrolysis

2. Detection of the Rotation of  Subunit H. Noji et al., 1997, Nature Biotinylation and fluorescent label of actin Assembly of  subunit and actin by streptavidin  4 binding sites for biotin Detection  subunit rotation under a fluorescence microscope in the presence of ATP

3. Stoichiometries of O 2 Consumption and ATP Synthesis xADP + xPi +1/2O 2 + H + + NADH  xATP + H 2 O + NAD +  X : P/O ratio or P/2e - ratio Before the chemiosmotic model  P/O is an integer  3 for NADH and 2 for succinate After the chemiosmotic model  No requirement for P/O to be integer  2.5 for NADH and 1.5 for succinate  Proton efflux : 10 for NADH, 6 for succinate  4 protons for 1 ATP synthesis

4. The proton-motive force for active transport Adenine nucleotide translocase  Antiporter  Transport ADP inside & ATP out side ( 4 - charges out & 3 + charges in)  by transmembrane electrochemical gradient (pmf) Phosphate translocase  Symporter  H 2 PO 4 & H +  Favored by transmembrane proton gradient ATP synthasome  ATP synthase + both translocases

5. NADH Shuttle Systems from Cytosol to Mitochondria Malate-aspartate shuttle  Liver, kidney, heart  2.5ATP/1NADH

6. NADH Shuttle Systems from Cytosol to Mitochondria Glycerol 3-phosphate shuttle  Skeletal muscle and brain  Electron transfer to Q  Complex III  1.5 ATP/ 1 NADH

7. 19.3 Regulation of oxidative phosphorylation

8. Regulation of Oxidative Phosphorylation Acceptor control of respiration  ADP (as a P i acceptor) Mass-action ratio  [ATP]/ ([ADP][P i ]); Normally high Inhibitory protein  IF 1 : bind to two ATPases as a dimer  Inhibition of ATP hydrolysis during hypoxic conditions (heart attack or stroke)  Dimerization is favored at low pH  Oxygen limitation  fermentation  lowering pH

9. Hypoxia Condition favorable for ROS generation Defense systems in Mito - SOD & Glutathione peroxidase - Regulation of PDH by PDH kinase - Replacement of one subunit of complex IV (COX4-1  COX4-2)

10. ATP-Producing Pathways are Coordinately Regulated Ratio of [ATP]/[ADP]  Coordinated regulation of major catabolic pathways

11. 19.4 Mitochondria in thermogenesis, steroid synthesis, and apoptosis Other important functions of mitochondria 1.Heat generation 2.Steroid hormone synthesis 3.Apoptosis

12. Heat generation by uncoupled mitochondria Thermogenin (uncoupling protein)  Unique protein in the mitochondria of brown adipocytes (brown adipose tissue; BAT)  Path for H + to matrix  Oxidation energy  Heat dissipation  Body temperature maintenance  Newborn mammals and hibernating animals

13. Mitochondrial P-450 Oxygenase Mitochondria is a site for steroid H production ; sex H, glucocorticoids, mineralocorticoids, Vit D H Adrenal glands & gonads Mitochondrial cytochrome P-450  Heme containing enz  Serial hydroxylation of cholesterol or related sterol R-H + O 2 + NADPH  R-OH + H 2 O + NADP +  Dozens of cyt P-450 located in the inner mito membrane  Complex e - flow from NADPH to P-450 heme ER cytochrome P-450 in hepatocytes  Similar catalytic mechanism as mito cyt P-450  Hydroxylation of xenobiotics  clearance through kidneys

14. Initiation of apoptosis Apoptosis (programmed cell death)  Critical during embryonic development  Conservation of cell’s molecular components  Triggered by external death signals or internal events (DNA damage, viral infection, ROS, stresses)  Mitochondrial cyt C

15. 19.5 Mitochondrial Genes: Their Origin and the Effects of Mutations

16. Mitochondrial Genome Circular, double stranded DNA ~ 5 copies of the genome in each mitochondria Human mitochondrial chromosome  37 genes (16,569 bp)  13 genes for proteins in respiratory chain  Genes for rRNA, tRNA for protein-synthesizing machinery

17. Origin of mitochondria Hypothesis of endosymbiotic origin  mt DNA, ribosomes, tRNAs  Enzymatic machinery for oxidative phosphorylation  F o F 1 complexes Respiration-linked H + extrusion (pmf)  Nutrient symport with H + (lactose)  Rotatory motion of bacterial flagella “Mitochondria is originated from endosymbiotic bacteria in primitive eukaryotes”

18. Mutations in mt DNA Accumulation of mt DNA mutation throughout the life  Major site for ROS  Ineffective systems for mistake correction during replication & Ineffective DNA-damage repair systems  Variation among individual cells in one organism & variation among organism

19. Photosynthesis: Harvesting light energy CO 2 + H 2 O light O 2 + (CH 2 O)

20. 19.6 General features of photophosphorylation

21. Photosynthesis in plants Two processes  Light reaction - light energy  ATP & NADPH, - O 2 release from  Carbon-assimilation reations (carbon-fixation) - CO 2  carbohydrate; using ATP & NADPH

22. Chloroplasts The light-dependent & the carbon-assimilation reactions  Outer & inner membranes  Thylakoids (arranged in stacks ‘grana’) ; its membrane embedding - Photosynthetic pigments - Enzyme complexes for light reactions & ATP synthesis  Stroma - Enzymes for carbon-assimilation reactions