Mitochondrium – Peroxisome - Mitochondrium – Peroxisome - ChloroplastChloroplast

outer membraneinner membrane

crista matrix

Dr. habil. Kőhidai LászlóAssoc. Prof.

Dept. Genetics, Cell & ImmunobologySemmelweis University

2008

HistoryHistory

AltmannAltmann – describes Mch – describes Mch BendaBenda -- name „Mitochondrium” name „Mitochondrium”

was was given by him given by him WarburgWarburg - invetigated the - invetigated the

enzymes enzymes of respiratory of respiratory chain chain

Lehninger Lehninger – described oxydative – described oxydative phosphorylation phosphorylation

MorphologyMorphology

Network of Mch in a Network of Mch in a fibroblast cell fibroblast cell

Detection of ATP-syntaseDetection of ATP-syntase

Characteristic dataCharacteristic data

• Size:Size: 7 x 0.5 7 x 0.5 mmBUT: wide range in different cell types !BUT: wide range in different cell types !

• Number: depends on the energy Number: depends on the energy requirements/budget of the cellrequirements/budget of the cell

sperim - 24sperim - 24WBC. - WBC. - 300300liver cell - 500-2500liver cell - 500-2500

Chaos-Chaos - Chaos-Chaos - 500.000 !500.000 ! amebaameba

CompositionCompositioncompartmentalisationcompartmentalisation

Outer membraneOuter membrane• poor in proteinspoor in proteins• characteristic protein: characteristic protein: porinporin• ((-sheet– trimers-sheet– trimers form channels)form channels)• permeability up to permeability up to 5000 dalton5000 dalton

Inner membraneInner membrane• 70% proteins70% proteins• ee-- - transporter chaini - transporter chaini• ATP synthesisATP synthesis• other point impermeable – 20% other point impermeable – 20% cardiolipincardiolipin

MatrixMatrix

• Pyruvate dehydrogenase complexPyruvate dehydrogenase complex• Enzymes of citric acid cycleEnzymes of citric acid cycle• Enzymes of Enzymes of -oxydation of fatty acids-oxydation of fatty acids• Enzymes of amino acid oxydationEnzymes of amino acid oxydation

• DNA, ribosomesDNA, ribosomes• ATP, ADP, PATP, ADP, Pii

• MgMg2+2+, Ca, Ca2+2+, K, K++

Inner membrane of MchInner membrane of Mch

cristacrista

tubulartubular

berry-likeberry-likefingerprint-likefingerprint-like

Localization in the cellLocalization in the cell

Basal striation

Mch as osmotic regulator of Mch as osmotic regulator of the cell the cell

Significant HSignificant H220 ration of matrix moves to 0 ration of matrix moves to the intermembraneous space and forms a the intermembraneous space and forms a „„condensedcondensed” comformation” comformation

normalnormal condensedcondensed

Relation of biochemical processes Relation of biochemical processes

in Mch.in Mch. pyruvate

fatty acid

Acethyl-CoA

Citric acid cycle

CO2

NADH+HFADH2

O2

H2O

ATP

Terms of Chemiosmotic Terms of Chemiosmotic theorytheory

• Mch. Respiratory chain – moves electronsMch. Respiratory chain – moves electrons - pumps H- pumps H++ into into

intermembrane spaceintermembrane space• Mch. ATP synthase works also as a HMch. ATP synthase works also as a H+ + pump.pump.

• Reversible mechanism:Reversible mechanism:

• Several carrier molecules for metabolites, ions – in the inner Several carrier molecules for metabolites, ions – in the inner membrane of Mch. membrane of Mch.

• Other point of the inner membrane of Mch. is impermeable for Other point of the inner membrane of Mch. is impermeable for HH++ and OH and OH--. .

H+ in ATP synthesis

ATP cleavageH+ out

I.I.

II.II.III.III. IV.IV.

UQUQ

H H ++ H H ++ H H ++

Matrix

Intermembranespace

I.I. NADH dehydrogenaseNADH dehydrogenaseII.II. Succinyl dehydrogenaseSuccinyl dehydrogenaseIII.III. Ubiquinone – cytochrom c oxydoreductaseUbiquinone – cytochrom c oxydoreductaseIV.IV. Cytochrom oxydaseCytochrom oxydase

Enzyme systems of Enzyme systems of inner membrane in Mchinner membrane in Mch

I.I.II.II. III.III.

IV.IV.

e e --

H H ++Acidic pHAcidic pHRedox potentialRedox potential INCREASING:INCREASING: I. < III. < IV.I. < III. < IV.

Matrix

Matrix

Intermembrane space

Intermembrane space

Resting phase

Ionophore treated (Valinomycin)

[H+]=10-9 M

[H+]=10-9 M

[H+]=10-9 M

[H+]=10-7 M

KK++

KK++

[K[K++] = [Cl] = [Cl--] = 0.1 M] = 0.1 M

[K[K++]<[Cl]<[Cl--]]

ATPH+

Electrochemical proton-Electrochemical proton-gradientgradient

pH gradientpH gradient membrane-potentialmembrane-potential

ATP synthesisATP synthesis

pH V

b-c1 complex

Q

cyt c

cytochrome oxydase

NADH dehydrogenase

OO22 HH22OO

NADHNADH NADNAD++

HH++

Electron transportElectron transportin Mch in Mch

Knob-like protusions of the Knob-like protusions of the inner Mch membraneinner Mch membrane

headhead

basisbasis

ATP-synthaseATP-synthase

proton carrierproton carrier

Structure of ATP-synthaseStructure of ATP-synthase

F1 ATP-ase(6 subunits)

Transmembrane H+ carriers (9 subunits)

ATP-synthaseATP-synthase

- rotor

a, b, - stator

Experimental evidenceExperimental evidence

H+

H+H+

H+

H+

H+H+

ADP + Pi

ATP

Bacterio-rhodopsin ATP synthase

ADP + Pi

ATP

ADP + Pi

ATP

H +

Transports required by Transports required by ATP-synthaseATP-synthase

Antiport Symport

Adenine nucleotie translocaseAdenine nucleotie translocase Phosphate translocasePhosphate translocase

H+

H+H+

H+

H2PO4-

H2PO

4-

ADP

ADP

ATP

ATP

I. III. IV.II.

H+ H+ H+H+

HeaHeatt

Brown adipose tissue Brown adipose tissue

MchMch.

thermogenin

TransportsTransportsSignal seq.

Receptor

Contact-pointTransloconGIP

! Hsp70 !! Hsp70 !

Mch. Hsp-kMch. Hsp-k

Origin of MitochondrionOrigin of Mitochondrion

• De novoDe novo synthesis synthesis• DivisionDivision

• Endosymbiont theoryEndosymbiont theoryArchaic Cyanobacteria – 1.5 x 109 yrs ago

porin (Gram (-) bact.)porin (Gram (-) bact.) electron transport chainelectron transport chain ATP synthaseATP synthase mt DNSmt DNS ribosomeribosome

BUT:BUT: GiardiaGiardia has NO Mch (anaerob) has NO Mch (anaerob)

Origin of Mitochondrion 2Origin of Mitochondrion 2

• Composition of outer membrane – Composition of outer membrane – eukaryotic type; the eukaryotic type; the inner membrane is inner membrane is composed by prokaryotic composed by prokaryotic componentscomponents

• Mch has own protein synthetic system, Mch has own protein synthetic system, the starter amino acid is the starter amino acid is formyl-Metformyl-Met

• Inhibitors of protein synthesis in Mch: Inhibitors of protein synthesis in Mch: antibioticsantibiotics acting on bacterial protein acting on bacterial protein synthesissynthesis

Network of Mch in budding Network of Mch in budding

S. cerevisiaeS. cerevisiae

IsotopeIsotopelabellinglabelling

CELL PROLIFERATIONCELL PROLIFERATION

mt-DNAmt-DNA

• ring shape, 5 –10 copies/Mch.ring shape, 5 –10 copies/Mch.• 20 Mch genes are coding proteins20 Mch genes are coding proteins• there are no intronsthere are no introns• few regulator genesfew regulator genes• no histonsno histons• repliation, transcription, translation repliation, transcription, translation • 22 tRNA, 2 rRNA22 tRNA, 2 rRNA

Human mt-DNAHuman mt-DNA

rRNArRNA

ND1; 2ND1; 2

I.I.II.II.ATP-syntaseATP-syntase

III.III.

ND3-6ND3-6

Cyt bCyt b

Mch myopathyMch myopathy

Singlefibre

Clusters of fibers

Crystallinestructurein the matrixof Mch

Mass ofpathologicMch-s

PeroxisomePeroxisome

•Single membrane coverageSingle membrane coverage

•Selective import of proteinsSelective import of proteins

•No genomeNo genome

•Oxydative enzymes:Oxydative enzymes:

catalasecatalase

urate oxydase urate oxydase (crystalloid)(crystalloid)

Origin of peroxisomeOrigin of peroxisome

• OO22 producing bacteria – early phase of phylogeny producing bacteria – early phase of phylogeny

• the Othe O22 is toxic to other cells/organisms is toxic to other cells/organisms

• peroxisomeperoxisome could neutralize the O could neutralize the O2 2 and its radicalsand its radicals in the cytoplasmin the cytoplasm

Functions of peroxisomeFunctions of peroxisome

•RHRH22 + O + O22 R + R + HH22OO2 2 (toxic) (toxic)

•HH22OO22 + R’H + R’H22 R’ + R’ + 2H2H22OO

catalase (liver, kidney)catalase (liver, kidney) -oxydation: alkyl chain - (C-oxydation: alkyl chain - (C22

ac.CoA)nac.CoA)n

Peroxisomes in plantsPeroxisomes in plants

• In plantsIn plants

leafs: leafs: photorespiration - Ophotorespiration - O22 consumption; COconsumption; CO2 2

germination: glyoxylate cycle germination: glyoxylate cycle (glyoxysome)(glyoxysome)

Fatty acidFatty acid ac. CoA succinate ac. CoA succinate glucoseglucose

Peroxisome in plantsPeroxisome in plants

glyoxisome

lipid

peroxisome

PeroxisomePeroxisome

• 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 syndromePXR1

ChloroplastChloroplast

Thylakoid membrane

(light reaction)

Stroma(dark reaction)

Oxygen-requiring bacteria move to regions where oxygen is being liberated by photosynthesis

Engelmann-experimentEngelmann-experiment(1894)(1894)

Photopigments of chloroplast

Z-scheme ofZ-scheme ofelectron transportelectron transport

in chloroplastin chloroplast

Chloroplast – NAPH / ATP synthesisChloroplast – NAPH / ATP synthesis

Comparison of ATP generation Comparison of ATP generation in Mch - Chloroplastin Mch - Chloroplast