8. Organelles: Mitochondria
Protein import into mitochondria
- Fully translated in cytosol - Bound by chaperone Hsp 70, prevents folding - Imported into mitochondria: -- Mitochondrial signal sequence -- post translational translocation
How does Electrochemical proton gradient drive ATP synthase?
- Protons flow down gradient through ATP synthase - This rotates ATP synthase (converted to mechanical energy) - Mechanical energy drives ATP synthesis from ADP + Pi
Where is ATP synthase located?
ATP synthase — embedded in inner mito membrane
Pyruvate & fatty acid broken down to what in the mitochondria?
Acetyl CoA
What is Acetyl CoA metabolized by?
Acetyl CoA metabolised by TCA reducing NAD+ to NADH
What protects from Reactive Oxygen Species damage?
Antioxidants
Electrochemical proton gradient
As electrons move along respiratory chain, energy is stored as electrochemical proton gradient across inner membrane Drives ATP synthesis
Pro-apoptotic bcl-2 proteins
BAD, BAX
simple summary of intrinsic pathway of apoptosis
BAX translocate into mito mebrane as a respinse to cell damage/stress BAX create pores in mito membrane Cytochrome C is released through these pores Cytochrome C activates caspase cascade apoptosis occurs
What is essential for inner membrane Impermeability?
Cardiolipin
What occurs in the outer membrane?
Cardiolipin synthesis Lipid modification
Mitochondria: Major functions
Cell's power plants ATP production (Aerobic respiration) Also; Cell signaling & apoptosis
Mitochondrial pore formation & Apoptosis steps
Pro-apoptotic bcl-2 proteins (BAD, BAX) translocate from cytosol to mitochondrial membrane in response to cell damage/stress & signalling ↓ Formation of pores in outer membrane (BAX) ↓ Release pro-apoptotic factors (cytochrome C) into cytosol ↓ Activation caspase cascade ↓ Apoptosis
Glycolysis & Mitochondria
Pyruvate (from glycolysis) & fatty acids enter mitochondrion & are broken down to Acetyl CoA in matrix
Reactive Oxygen Species can inactivate what? what is the result of the inactivation?
ROS can inactivate ETC e- acceptors: further build-up of ROS
What diseases are Reactive Oxygen Species implicated in?
ROS implicated in degenerative diseases(Alzheimer), cancer & aging
Shape of mitochondria in skeletal muscle
Skeletal muscle: rows, nestled between myofibrils
Shape of mitochondria in sperm
Sperm: wrapped around flagellum
Summary of Energy-producing reactions of the Mitochondrion
TCA cycle (creates CO2 + NADH) Electron transport chain uses energised electrons from NADH to pump protons into intermembrane space Protons fall down electrochemical gradient through ATP synthase, making ATP Inner membrane very impermeable to hold ion gradient
Import proteins of the mitochondria
TOM & TIM
Electron transport & ATP synthesis: summary
Transfer of electrons to electron acceptors in the inner membrane results in pumping of protons out of mitochondrial matrix to the intermembrane space, which return to power ATP synthase ultimately ½ O2 will take e- and become water
TIM
Translocase of Inner Membrane requires electrochemical gradient
TOM
Translocase of Outer Membrane - Hsp70+protein binds to TOM - TOM recognises mt signal sequence
What is driven by Mitochondrial gradient?
Transport processes driven by the gradient used to bring essential components in and export essential products out
Shape of mitochondria in Steroid secreting cells
Tubular cristae
What does Thermogenin (uncoupling protein) uncouple?
Uncouples respiration from ATP synthesis
Bcl-2 family
can be pro or anti apoptotic Bcl-2 is blocking release of cytochrome C
What protects peroxisomes from Reactive Oxygen Species?
catalase
What complexes are involved in Electron transport ?
complex I complex III complex VI
What does the TCA (citric acid/Krebs) cycle produce?
creates CO2 + NADH
Protons flow (up/down) gradient through ATP synthase
down gradient
What occurs in the matrix?
enzymes of oxidative metabolism - Pyruvate oxidation - Fatty acid oxidation - TCA cycle (Citric acid cycle; krebs cycle)
Apoptosis: Extrensic pathway
extrinsic pathway == death ligand and receptor
two Antioxidants?
glutathione peroxidase superoxide dismutase
Cardiolipin synthesis
super important for inner membrane but actually made in outer membrane
Mitochondria protein import
the Mitochondria is not self sufficient - most proteins must be imported in - Majority of proteins encoded by nuclear genes
Thermogenin
uncoupling protein allows protons to "leak" from cytoplasm to matrix uncouples gradient: uncoupling —> bypassing ATP synthesis, so no making of ATP
What is driven by the voltage gradient?
voltage gradient drives ADP-ATP exchange
oxidative phosphorylation
— making of ATP — in the presence of Oxygen (no other way)
Caspase cascade
→ cell death in both pathways, the enzyme that actually mediates apoptosis
Function of ATP synthase
Couples oxidation to phosphorylation mechanical rotation leads to phosphorylation energy transformation from mechanical to biochemical energy Transmembrane H+ carrier Head: ATPase Subunits bind to ADP + Pi (in the matrix)
Function of BAX
Formation of pores in outer membrane; which allows release of pro-apoptotic factors (cytochrome C)
What can diffuse across the outer membrane?
Free diffusion of small molecules & ions
Mitochondrial Proteins are made where?
Fully translated in cytosol
Steps of protein import into Mitochondria
1. Hsp70+protein binds to TOM; TOM recognizes mito signal sequence 3. Precursor translocated across inner membrane via TIM; requires electrochemical gradient 4. Signal sequence removed by matrix protease 5. Protein spontaneously folds or helped by chaperone Hsp60 & ATP
Pyruvate oxidation
2 from splitting of glucose
Cardiolipin
20% inner membrane lipid "double phospholipid"; 4 tails; two glycerol backbone Made in mitochondria outer membrane, most others imported Impermeability to ions
Shape of mitochondria in hepatocytes
Hepatocyte: oval with numerous cristae
Mitochondria: Shape & distribution
Lamellar cristae: MOST cells
The Energy released from what used to generate ATP?
oxidation of nutrients Oxidative phosphorylation
What is driven by the pH gradient?
pH gradient drives pyruvate import and phosphate import
Mitochondria: Outer membrane
permeable to small molecules & ions (~5-10 kDa) biochemical reactions- primarily lipids (made)
What kind of translocation occurs?
post translational translocation
Cytochrome C
pro-apoptotic factors if Cytochrome C is released from mito — cell undergoes apoptosis
NAFH+
proton donor in ETC High energy electrons from NADH passed along electron transport chain to O2
What does the Electron transport generate?
proton gradient across inner membrane → drives ATP synthesis
What can Reactive Oxygen Species damage?
Damages protein, RNA, DNA (mtDNA: no histones & ↓ DNA repair)
How do mitochondria divide?
Divide by binary fission Replicate DNA & divide in response to energy needs of cell (not linked to cell cycle) so if you need more energy, mito will divide
the mother of all reductions
ETC (electron donation) —> O2 to H2O
What occurs in the inner membrane?
ETC & ATP synthase e- transport chain oxidative phosphorylation (ADP—>ATP)
What import complex is is the inner membrane?
ETC complexes
What drives ATP synthasis?
Electrochemical proton gradient
Mitochondria: Inner membrane
Impermeable to most molecules Cristae ↑ surface area ETC ATP synthase***
Outer membrane has what kind of receptors?
Import receptors for larger proteins
Inner membrane has what kind of receptors?
Import receptors for matrix proteins
What is the energy conversion catalysed by the mitochondrion?
Inner membrane changes chemical bond energy (NADH oxidation) into phosphate bond energy (in ATP) oxidative phosphorylation
Apoptosis: 2 different pathways
Intrinsic & extrinsic pathways
Is mitochondrial division related to the cell cycle?
NO; if you need more energy mito will divide
Mitochondria: Localization & Numbers
Near sites of high ATP requirement # varies in diff. tissues (1 - several thousand) Numbers & energy producing enzymes (TCA & ETC) increase with need (ie: regular exercise) example: tail of sperm, mito surround flagella
What produces heat in mito?
Oxidative metabolism (TCA, ETC) produces HEAT
Barth syndrome
X-linked cardiolipin synthesis disorder inner membrane becomes permeable Cardiomyopathy Generalised muscle weakness & chronic fatigue Neutropenia High mortality in infancy: Sudden infant death Infection Cardiac failure
Inner membrane permeability
impermeable to small, charged molecules
Brown adipose tissue
in newborns and babies fat with many specialised mitochondria uncoupling protein (UCP); thermogenin, in inner membrane forms channels through membrane
Where does glycolysis occur?
in the cytoplasm
Where is the site of ATP synthase?
inner membrane
Porins
integral proteins found in the outer membrane
In the inter membrane space the concentration of H+ ions is _______; so the pH is _______.
inter membrane space concentration of H+ ions is high, pH is low
Apoptosis: Intrinsic pathway
intrinsic pathway == think mitochondria **
Besides energy, what else is produced by oxidative phosphorylation in the mitochondria?
most endogenous Reactive Oxygen Species (ROS)
Mitochondria: Matrix
mtDNA mt ribosomes, tRNA, rRNA oxidative metabolism: production of ATP