Mitochondria Biology PDF

Chapter 12 Mitochondria, (Chloroplasts), and Peroxisomes. 8th ed. Chap 3 9th ed. Chap12 Metabolism 85-97 Mitochondria 411- Glycolysis and Basic structure, citric acid cycle Evolution Electron transport Protein Import...

Chapter 12 Mitochondria, (Chloroplasts), and Peroxisomes. 8th ed. Chap 3 9th ed. Chap12 Metabolism 85-97 Mitochondria 411- Glycolysis and Basic structure, citric acid cycle Evolution Electron transport Protein Import ATP synthesis Glycolysis and citric acid cycle (not 8th ed. Chap13 detailed) Mitochondria 425- Electron transport 437 ATP synthesis Basic structure, Chloroplasts and Evolution other plastids 458 Protein Import Peroxisome (brief) Chloroplasts and 451 other plastids 458 Peroxisome (brief) 445  Mitochondria basics Majority of ATP is produced here 1) Two membranes = Inner and outer membrane 2) Two spaces = Matrix and intermembrane space.  Mitochondrion, basics 1) Inner membrane has Electron transport system to make H+ gradient across inner membrane, ATP synthase (F1F0- ATPase), Transporters to incorporate metabolites, 2) Inner membrane isADP/ATP and Pi etc, full of proteins, but not rigid. Mt is flexible and continuously moving, 3) Inner membrane needs dividing and fusing. large surface area, so it is folded into Cristae. 4) Matrix (inside of inner membrane) contains enzymes for citric acid cycle, b-oxidation, and mitochondrial DNA, ribosome, replication enzymes etc. 5) Outer membrane has porin: free pass for molecules smaller than ~1000 daltons; Amino acid, pyruvate, fatty acid and glycerol are small enough to go through it. Porin ATP synthesis Food  ATP  Digestive Starch er) tract s e p o lym (gluco Amylase Blood Absorption from intestinal lumen to blood stream “Blood sugar” Intake to cell Cell Mitchondria. Glycolysis Glucose Pyruvate 2 ATP ~29 ATP  Glycolysis (in cytosol) In Glycolysis (Glucose  2 pyruvates) 2 x ATP are produced per glucose. 2 x NADH are produced per glucose. NAD = Nicotinamide adenine dinucleotide et a ils. d emorize ) e ed to m is class No n st for th a ( a t le NAD = Nicotinamide adenine dinucleotide High energy reduction NAD+ NADH glycolysis In mitochondrial matrix Pyruvate Acetyl-CoA Coenzyme-A Citric acid After Glycolysis cycle Acetyl group of Pyruvate is (TCA cycle) transferred to CoA, to form acetyl-CoA. Citric acid cycle oxidizes all carbon to CO2 4xNADH, 1xFADH2 and 1xGTP, are produced per one pyruvate.  Glycolysis (cytosol) one glucose -> 2 pyruvate Produces 2 ATP & 2 NADH (in cytosol) TCA cycle + pyruvate dehydrogenase (in mitochondria) 2 pyruvate -> 6 CO2. Produces 2 GTP (=2ATP) 8 NADH Sub Total: 2 FADH2 one glucose  6 CO2 (C6H12O6 +6H2O  6CO2 + + 24H) 4 ATP 10 NADH Majority of ATP is produced from them 2 FADH2 by Oxidative phosphorylation. 1. Electron transport system 2. ATP synthase  Cytosolic NADH needs extra-energy to get into Mt. Matrix 2 X NADH 8 Outer NADH membrane Inter- membrane Space (Biochemical detail) NADH is impermeable: Converted to malate, then antiport with ketoglutarate and then reduced to produce NADH again.… (Malate-aspartate shuttle), ~~60% energy yield.  Electron transport reductio n High energy H- Oxidizati on NAD+ NADH Redox potential How energetically favorable to reduce (give electron to) other chemical. = strength as an electron donor Pump H+ Bang! 2e- 2e- 2e- NAD NADH+ Ubiquinone Oxidized Reduced Oxidized Reduced Oxidized (low energy) (high (low energy) energy) (high energy) (low energy)  NADH dehydrogenase complex (Complex I) Pump H+ Bang! 2e- 2e- 2e- NAD NADH+ QH Q Ubiquinone 2 Oxidized Reduced Oxidized Reduced Oxidized (low energy) (high (low energy) energy) (high energy) (low energy) NAD+ NAD  2 e- NAD+ NADH H+ H+ Bang! 2e- 2e- NAD+ NADH Oxidized Reduced (low energy) (high energy)  Electron transport Ubiquinone (Co-Q) can receive two electrons with two protons. Cytochrome C can bind an electron.  H2O Complex II does not pump Stoichiometry of H+ pump: H+. 1)Electron pair from NADH pass through 4 H+ by complex I three complexes, which pump up H+ 4 H+ by complex III out of Matrix. 4 H+ by complex IV 2)Electron pair finally reduces O2 and produce H2O by Complex IV. Total 12 H+ per NADH 3)FADH2 has lower redox potential than 8 H+ per FADH2 NADH: Complex II receives electrons  ATP Synthase (F1F0 ATPase) 1. Final step of ATP synthesis 2. Convert electrochemical gradient of H+ into ATP 4 4 4 (1978)  Chemiosmotic theory  Free energy created by H+ electrochemical gradient is given  That energy can be used for ATP generation. = It is not metabolic ATP synthesis = Closed membrane is essential for the reaction (like a dam) = pH change affect ATP synthesis by isolated Mt. Everything make sense but still the idea looked quite  ATP Synthase (F1F0 ATPase) F0: 10-14 c-subunits 1 a-subunit F0 2 b-subunits c-ring = ring of c- subunits Stator stalk = two b subunits. g F0 = transmembrane H+ carrier. F1 F :a ba bthe 1 gde 3 3 spherical structure. 3 3 gde the rotor stalk (g-stalk). F1 = Catalytic component of ATP synthesis. matrix  Structure of F1F0 ATPase and blender. g Energy (H+ gradient or electric power) is first converted to the rotation of rotor, and the rotation causes the structural change of protein inside the vessel to perform chemical reaction (ATP synthesis or crashing apples). 1) While H+ pass through F0, the rotor rotates relative to the stator. in F0: H+ Gradient  rotation 2) While rotor rotates, F1 synthesize ATP from ADP and phosphate. in F1: rotation  ATP synthesis  H+ 1) a-subunit has two half-channels How it rotates for H+. H+ 2) Each c-subunit has a proton- acceptor, which receives proton from a half channel. 3) After accepting proton, c-ring slides by one subunit. Then next H+ comes through the half channel and binds next c- subunit. Then, c-ring rotates H++ by one subunit… H++ H H H+ + + 4) After one round of rotation, H+ is transferred from c-subunit to another half-channel that is open to the opposite side of the membrane. Rotation g 10-14 H+/turn Intermembra This class assume that one c- ne space ring has 12 c-subunits.  How rotation of g-stalk makes ATP in F1 Open 1) (ab)3 g Tight Low g-stalk (ab)3 complex  g-stalk rotates with c-ring.  (ab)3 complex in F1 is fixed by stator stalk (b-subunit), so it does not rotate with g-stalk.  As g-stalk rotates in F1 ATPase, three ab subunits change conformation.  Open O (open)-conformation has low affinity to (ab)3 ATP and can exchange ATP/ADP. g T (tight)-conformation can bind only Low Tight ATP, never release it. 120º turn L (low affinity)-conformation binds to Low ADP and Pi but they can’t escape from the protein. Open g Tight Open conformation released ATP, then binds ADP & Pi Low g Open Tight T (tight)-conformation can convert ADP +Pi to ATP. 120º turn Tight 3) Open g Low Every 120o trun, one ATP is produced. Tight Therefore, 3 ATP are produced per one g Open complete turn of g-stalk in F1 Low ATPase 120º turn Open 4) g Low Tight  Stoichiometry.. 3 ATP As g-stalk rotates one turn, theoretically, ab complex produce 3 ATPs per turn. 12 (10 – 14) H+ are consumed per turn, because a c-ring has 12 (10- 14) subunits.  Assuming that c-ring has 12 c- subunits, ~12H+/3 ATP = 4 4 H+ can produce one ATP. Remember that: electron transport pumps 12 H+ 12 H+ / NADH 8 H+ / FADH2 One matrix NADH  12 H+  3 ATP One cytosolic NADH (60% efficient)  ~7.2 H+  1.8 ATP One FADH2  8 H+  2 ATP Grand total, ATP per glucose ATP H+ pumped produced x12x0.6 ~14 by F1F0 x12 1/4 31.5 96 126 H+ ATP x8 16 Glu + 6O2  6H2O + 6 CO2 (-2780 kJ/mol) ATP + H2O  ADP + Pi (-30 kJ/mol) 30 kJ/mol x 35.5 molecules = 38.3% 2780 kJ/mol However, H+ gradient is also used for many coupled transport systems.  Transport of metabolites across the inner membrane  Many transport systems utilize pH gradient, product/substrate exchange.  So, H+ gradient is not used only for ATP synthesis. ATP synthase reaction is reversible ATP synthase can hydrolyze ATP to produce H+ gradient by the reverse reaction of ATP synthesis. Here, ATP hydrolysis by F1 ATPase turn the rotor, so that proton is pumped out.  BioPhysicists’ Toy: ATP-driven energy-efficient motor. g Noji, H. et al., Nature 386, ~10 nm 299-302 (1997). Smallest motor: Nearly 100% energy efficiency  Mitochondria genome (16.6 kbp) NADH NADH dehydrogenase dehydrogenase subunits subunits Figure 14-58, Mol Biol. Cell, 4th. MT has DNA. Some mitochondrial protein are encoded in MT genome. E.g. 7 out of 42 subunits of complex I. MT contains 1000 to 1500 different proteins: majority are encoded in nuclear genome. Human mitochondria has 37 genes, 13 of which are encoding proteins. All MT tRNA are encoded in MT genome  aerobic Endosymbiont Hypothesis Prokaryote ~ 2 billion years ago Complete genome for early MT Early anaerobic Early aerobic Eukaryote (Endosymbiosis) Eukaryote Transfer DNA from MT to 1. Aerobic prokaryote (which chromosome. has electron transport and ATP synthase system) might be engulfed to early Loses different anaerobic eukaryote to set of genes. become first MT. 2. Then some MT genes were transferred to nucleus during evolution of eukaryotes. animals Fungi Plants, etc (Some of) supporting evidence of Endosymbiont hypothesis 1. MT has its own DNA and replication machinery. 2. MT has its own ribosomes rRNA, tRNA, RNA polymerase etc. 3. These proteins are homologous to prokaryotes, thus useless for nuclear genome and nuclear gene expressions. 4. MT divides like bacteria (no spindle, no cell cycle-phase), and not regulated by host cell cycle (mostly). 5. MT DNA is circular and attached to the inner membrane, and replicate like bacterial chromosomal DNA. Fusio Mitochondrial division and fusion (only n in 9th ed.) Divisio For fusion, Protein- n protein interaction and GTP hydrolysis is involved. as. l tai de e th r ize o em m to ed ne No For division, dynamin-related protein-1(Drp1) pinches off the membrane, by hydrolyzing GTP.  Control of Mitochondria Mitochondria behave (somewhat) like independent microbes in cytoplasm. But there are some regulations. Metabolic regulation Number per cell Localization Tissue-specific functions: eg. steroid synthesis in adrenal cells heme biosynthesis in bone marrow As semi-independent bug, MT has no vesicular traffic from/to ER, Golgi, or plasma membrane. They are not connected with cytoskeleton (usually). How they get their protein and lipid?  Protein import Majority of MT proteins are translated by cytosolic ribosome. They are transported to MT after translation is done. N-terminal 20 to 55-aa sequence (presequence) serve as a signal for matrix and inner membrane. John Tackett, Age 15 4 different destinations: Matrix, Inner membrane, Intermembrane space, Outer membrane Two translocators: Outer membrane translocator (TOM) Inner membrane translocator (TIM)  1. Cytosolic Hsp70 Protein import into unfold the protein matrix 3. ATP hydrolysis by hsp70 pushes protein 2. Tom complex in Tom. recognizes signal (presequences) John Tackett, Age 15 4. Tom-Tim interacts 6. Some are released as inner 5. Negative charge membrane attracts the proteins presequence 8. ATP hydrolysis 7. Presequence is cleaved, and and release of MT MT Hsp70 pull the protein into Hsp70 matrix  Protein import into mitochondria (other passways)  Multi-pass transmembrane proteins have internal import signals and stop transfer signals, and recognized by other Tims.  Inner membrane proteins that are encoded on MT genome are translated by MT ribosome and transferred to other translocase.  Transport to outer membrane or intermembrane space are also through the Tom complex, then laterally released, or once released to intermembrane space and then incorporated to outer membrane. No n ee d to m e m or ize th is slid e Lipid is delivered from ER to MT by protein MT does not have ability to synthesize its membrane lipid.