outer membrane inner membrane crista matrix Mitochondrium Peroxisome
outer membrane inner membrane crista matrix Mitochondrium – Peroxisome - Chloroplast Dr. habil. Kőhidai László Assoc. Prof. Dept. Genetics, Cell & Immunobology Semmelweis University 2008
History v Altmann – describes Mch v Benda - name „Mitochondrium” was given by him v Warburg - invetigated the enzymes of respiratory chain v Lehninger – described oxydative phosphorylation
Morphology
Network of Mch in a fibroblast cell Detection of ATP-syntase
Characteristic data • Size: 7 x 0. 5 mm BUT: wide range in different cell types ! • Number: depends on the energy requirements/budget of the cell sperim - 24 WBC. 300 liver cell - 500 -2500 Chaos-Chaos - 500. 000 ! ameba
Composition compartmentalisation Outer membrane • poor in proteins • characteristic protein: porin • (b-sheet– trimers form channels) • permeability up to 5000 dalton Inner membrane • 70% proteins • e- - transporter chaini • ATP synthesis • other point impermeable – 20% cardiolipin
Matrix • Pyruvate dehydrogenase complex • Enzymes of citric acid cycle • Enzymes of b-oxydation of fatty acids • Enzymes of amino acid oxydation • DNA, ribosomes • ATP, ADP, Pi • Mg 2+, Ca 2+, K+
Inner membrane of Mch crista tubular fingerprint-like berry-like
Localization in the cell Basal striation
Mch as osmotic regulator of the cell normal condensed Significant H 20 ration of matrix moves to the intermembraneous space and forms a „condensed” comformation
Relation of biochemical processes in Mch. pyruvate fatty acid Acethyl-Co. A CO 2 ATP Citric acid cycle O 2 H 2 O NADH+H FADH 2
Terms of Chemiosmotic theory • • • Mch. Respiratory chain – moves electrons - pumps H+ into intermembrane space Mch. ATP synthase works also as a H+ pump. H+ in Reversible mechanism: H+ out ATP synthesis ATP cleavage • Several carrier molecules for metabolites, ions – in the inner membrane of Mch. • Other point of the inner membrane of Mch. is impermeable for H+ and OH-.
H+ Intermembrane space H+ H+ UQ I. II. IV. Matrix I. III. IV. NADH dehydrogenase Succinyl dehydrogenase Ubiquinone – cytochrom c oxydoreductase Cytochrom oxydase
Enzyme systems of inner membrane in Mch I. Acidic p. H Redox potential INCREASING: I. < III. < IV. II. H+ e- III. IV.
Resting phase Matrix [H+]=10 -9 M [K+] = [Cl-] = 0. 1 M [H+]=10 -9 M Intermembrane space Matrix [K+]<[Cl-] [H+]=10 -9 M ATP H+ K+ Intermembrane K+ space [H+]=10 -7 M Ionophore treated (Valinomycin)
Electrochemical proton-gradient p. H gradient Dp. H membrane-potential DV ATP synthesis
NADH NAD+ NADH dehydrogenase H+ Q b-c 1 complex Electron transport in Mch cyt c cytochrome oxydase O 2 H 2 O
Knob-like protusions of the inner Mch membrane ATP-synthase proton carrier head basis
Structure of ATP-synthase F 1 ATP-ase (6 subunits) Transmembrane H+ carriers (9 subunits)
ATP-synthase e - rotor a, b, d - stator
Experimental evidence
Bacterio-rhodopsin ATP synthase H+ ADP + Pi H+ ATP H+ H+ + H H+ H+
ATP ADP + Pi H+ ADP + Pi ATP
Transports required by ATP-synthase Symport Antiport H+ ATP H 2 PO 4 - H+ ADP ATP ADP H+ Adenine nucleotie translocase H 2 PO 4 - H+ Phosphate translocase
Brown adipose tissue Mch. H+ I. II. H+ III. H+ Heat H+ IV. thermogenin
Transports Signal seq. ! Hsp 70 ! Receptor Mch. Hsp-k Contact-point Translocon GIP
Origin of Mitochondrion • De novo synthesis • Division • Endosymbiont theory Archaic Cyanobacteria – 1. 5 x 109 yrs ago § porin (Gram (-) bact. ) § electron transport chain § ATP synthase § mt DNS § ribosome BUT: Giardia has NO Mch (anaerob)
Origin of Mitochondrion 2 • Composition of outer membrane – eukaryotic type; the inner membrane is composed by prokaryotic components • Mch has own protein synthetic system, the starter amino acid is formyl-Met • Inhibitors of protein synthesis in Mch: antibiotics acting on bacterial protein synthesis
Network of Mch in budding S. cerevisiae
CELL PROLIFERATION Isotope labelling
mt-DNA • ring shape, 5 – 10 copies/Mch. • 20 Mch genes are coding proteins • there are no introns • few regulator genes • no histons • repliation, transcription, translation • 22 t. RNA, 2 r. RNA
Human mt-DNA r. RNA Cyt b ND 1; 2 ND 3 -6 I. III. ATP-syntase
Mch myopathy Single fibre Crystalline structure in the matrix of Mch Clusters of fibers Mass of pathologic Mch-s
Peroxisome • Single membrane coverage • Selective import of proteins • No genome • Oxydative enzymes: catalase urate oxydase (crystalloid)
Origin of peroxisome • O 2 producing bacteria – early phase of phylogeny • the O 2 is toxic to other cells/organisms • peroxisome could neutralize the O 2 and its radicals in the cytoplasm
Functions of peroxisome • RH 2 + O 2 • H 2 O 2 + R’H 2 R + H 2 O 2 (toxic) R’ + 2 H 2 O catalase (liver, kidney) · b-oxydation: alkyl chain - (C 2 ac. Co. A)n
Peroxisomes in plants • In plants leafs: photorespiration - O 2 consumption; CO 2 germination: glyoxylate cycle (glyoxysome) Fatty acid ac. Co. A succinate glucose
Peroxisome in plants peroxisome glyoxisome lipid
Peroxisome • import of proteins - 3 amino acid signal sequence on C-terminal - PAF-1 – peroxisomal assembly factor-1 PAF-1 • Zellweger syndrome protein to be impoted is affected - empty peroxisomes (brain, liver, kidney affected; lethal)
Gene transfection Zellweger syndrome PXR 1
Chloroplast
Thylakoid membrane (light reaction) Stroma (dark reaction)
Engelmann-experiment (1894) Oxygen-requiring bacteria move to regions where oxygen is being liberated by photosynthesis
Photopigments of chloroplast
Z-scheme of electron transport in chloroplast
Chloroplast – NAPH / ATP synthesis
Comparison of ATP generation in Mch - Chloroplast
- Slides: 50