Cell and molecular biology :MITOCHONDRIA AND RIBOSOME

1. MITOCHONDRIA & RIBOSOME
AMRUTHA K H
II MSc ZOOLOGY

2. MITOCHONDRIA

3. • Granular or filamentous organelles
• Seen in cytoplasm of plant and animal cell
• Absent in RBC of mammals and prokaryotic cell
• In prokaryotes ,located on the plasma membrane
• Perform series of biochemical and functional
properties
• Contain large battery of enzyme and co-enzymes
• Energy transducing system, recover energy and
convert into ATP.Hence called “power plant of
cell”

4. • Energy released by combustion with oxygen-aerobic
respiration
• In cells energy transformation takes place by 2 transducing
systems, mitochondria & chloroplast
• Main function of chloroplast is photosynthesis(endergonic)
and that of mitochondria is oxidative
phosphorylation(exergonic)

5. Number and Distribution
• Varies in different organisms
• Micromonas-1 mitochondrion,yeast-10,100
to 1000 in vertebrate cells
• In liver 1000-1600 mitochondria per cell
• Highest no: in flight muscles of certain
insects
• Uniformly distributed in cytoplasm
• In kidney tubules they are related to plasma
membrane for supply of energy

6. Shape & Size
• Typically sausage shaped
• May vary from granular,filamentous,club-shaped
or ring like to swollen and round shapes
• Size is large enough to be seen by light
microscope
• Width 0.5µ
• Length varies from 1.5µ to 7µ
• Smallest –yeast cell
• Largest- oocyte of amphibian

7. Gross structure
• Double walled, outer and inner membrane,
enclose compartment
• At certain points outer membrane is connected
to ER
• Outer membrane encloses outer chamber and
inner membrane encloses inner chamber
• Infolding of inner membrane-cristae
• Outer chamber has fluid of low viscosity
• Inner chamber contains dense proteinaceous
material-mitochondrial matrix

9. • Outer surface of inner membrane-C face
• Inner surface facing matrix –M face
• Mitoblast-mitochondria from which outer
membrane is stripped off
• Structural variation due to variation in cristae
• No: of cristae varies
• Fewer cristae in liver cells, numerous in muscle
cells
• Correlation exist between no: of cristae and
oxidative activity of cell

10. • 2 conformational stage for mitochondria :
1. Orthodox stage-ADP low, matrix more, cristae
well defines and no phosphorylation
2. Contracted stage-ADP more, matrix dense,
cristae not clear and phosphorylation
• Crests are studded on M side with particles-F1
particle
• Becomes visible only by negative staining.F1
particle has a base piece, stalk and head piece
• Presence of F1 gives symmetry to inner
membrane, related to direction of proton pump

11. • In choloroplast,F1 particles are on the outer side
• Mitochondrial matrix is usually homogenous,
may contain some ribosome,cDNA,protein and
host of other molecules

12. Chemical Composition
• 73% proteins
• 25 to 30% lipid of which 90% are phospholipids
and remaining cholesrol,carotenoids,vit E,free
fatty acids and triglycerides
• Outer membrane has 60% protein and 40% lipid
with large amount of cholesterol
• Matrix has compounds of sulphur,copper etc
• Battery of enzyme and small amount of DNA
and RNA.

13. Enzymes
• Definite compartmentalization for enzymes
• Outer membrane contains NADH-cytochrome-C-
reductase system, that contains flavoprotein and
cytochrome b5
• Most specific enzyme-monoamine oxidase
• Also contain kynurenine hydeoxylase,fatty acid
• Space between 2 membrane contains adenylate
kinase and nucleoside diphosphokinase

14. • Inner membrane contains respiratory enzymes,ATP
synthasee,succinate dehydrogenase,β-hydroxy
butyrate dehydrogenase and carinitine fatty acid
acyl transferase
• Matrix contain malate and isocitrate
dehydrogenase,fumarase,aconitase,citrate
synthetase,α-keto acid dehydrogenase and β-
oxidation enzyme
• Contains soluble enzymes of krebs cycle,DNA,RNA
and components of protein synthesis

15. Bioenergetics and Mitochondria
• Molecular orientation is complex
• Only needs entry of phosphate,ADP and acetyl
coenzyme A to produce ATP,C02,H20
• Energy liberated is used to synthesize new
molecules, for work etc
• 1st step in energy release is glycolysis.
• Under anaerobic condition, glucose is degraded
to lactase
• Achieved by 10 enzymes located in cytosol
C6H12O6 + 2 NAD+ + 2 ADP + 2 P ----->2C3H6O3+2ATP+2H2O

16. • under aerobic condition, products are pyruvate
and NADH.Since pyruvate contain large amount
of energy it further degrade.
• This occurs inside mitochondria by a series of
reaction called respiratory cycle.
• It is completed in 2 steps Krebs cycle and
oxidative phosphorylation.
• Krebs cycle takes place in mitochondrial matrix
• 1st step in the common pathway for degradation
of fuel molecules
• It undergoes 2 transformation stage

19. •Complex I- NADH dehydrogenase
•Complex II-Succinate dehydrogenase
•Complex III- Cytochrome C1 reductase
•Complex IV- Cytochrome C1 oxidase
•Complex V-ATP synthase

20. • ETC is coupled with the phosphorylating system at 3
points,
• In electron transfer, the protons are translocated
across the membrane from M side to C side.
• According to chemiosmotic hypothesis, this
translocation generates pH difference and a
membrane potential.
• These forms the proton-motive force, helps to move
H+ ions from C to M side
• It function as a proton pump
• 6 H+ ions on the M side give rise to 3ATP
NADH+H++3ADP+3Pi+1/2O2NAD++4H20+3ATP

21. • Phosphorylating system is represented as F1
ATPase.It has 3 parts :
1. Head piece
2. Hydrophobic protein
3. Stalk
• Energy balance of aerobic respiration shows
that 36ATP are produced from a glucose
C6H1206+36Pi+36ADP6CO2+36ATP+42H20

22. Mitochondrial DNA

23. • Circular in shape,5µm long
• Highly twisted ds molecule
• Can replicate and give rise to several circles
• Higher G-C content compared to nuclear DNA,
hence higher density
• MtDNA behaves as mitochondrial chromosome
• Replication of mtDNA and nuclear DNA takes
place at different time
• MtDNA duplicates during G2 phase of
cytokinesis.

24. Mitochondrial RNA
• 3 species of RNA are isolated-23S,16S and 4S
• They are synthesized on mtDNA template

25. Mitochondrial Ribosome
• Smaller than cytoplasmic ones
• Sedimentation coefficient of 55S with 35S
and 25S subunit
• More similar to bacterial ribosome
• Protein synthesis takes place in mitochondria
with the help of mitochondrial ribosome

26. Mitochondrion as Semiautonomous
Organelle
• Posses its own DNA,RNA and ribosome
• Capable of replication,transcrption and synthesis of
its own structural proteins
• Genetic code is different from the “universal code”
• RNA is resistant to action of cytoplasmic ribonuclease
• Time of replication is controlled by nuclear DNA.
• Formation of mitochondria is controlled by 2 system-
mitochondria and nuclear DNA

27. Functions
1. Respiration or oxidation of food stuffs
2. Transduction of energy
3. Synthesis of protein
4. Yolk formation
5. ATP transport
6. Formation of mitochondrial spiral
7. Lipid synthesis
8. Extra chromosomal inheritance
9. Heat production
10. Storage

28. RIBOSOME

29. • Submicroscopic particle
• Formed of ribobucleoprotein
• Facilitate protein synthesis, hence called
“protein factories” of the cell.
• 1st isolated by Albert Claude

30. • Siekevits and Zamecnik showed that protein
synthesis takes place in them
• R.B.Roberts named them ribosome due to
their RNA rich content

31. Occurrence and Distribution
• In both prokaryote and eukaryote
• In prokaryotic cell-freely in cytoplasm
• In eukaryotic cell-freely in cytoplasm or
attached to outer surface of ER-known as
cytoribosome
• Organellar ribosome-seen within organelles
like mitochondria and plastids

32. Number
• Cells active in protein synthesis have
numerous ribosome
• Many ribosome in cancer cells
• Mature mammalian erythrocyte have no
ribosome

33. Types
• According to size and sedimentation coefficient,2
types-70s and 80s
• 70s-found in prokaryotic cell
• 802 –found in eukaryotic cell

34. Structure
• Spherodial structure
• Porus,hydrates,2 subunit-one larger one smaller
• Smaller above larger as a cap like structure
• Separated by narrow cleft
• 70s-larger 50s+ smaller 30s
• 80s- larger 60s+ smaller 40s
• 2 subunits are united by Mg2+
• Below certain level of Mg2+,2 subunit separate-”first
critical level”
• This is reversible

35. • Further lowering below first critical level breaks
the 2 subunit-”second critical level”
• Normally 2 subunits remain free in cytoplasm, get
united only during protein synthesis.
• Mitochondrial ribosome occur in wide variety of
forms, most common is 55s ribosome. It contains
larger 40s and smaller 30s
• Mitoribosmes need more Mg2+ for stability

36. Ultrastructure-80s

37. • 60s appear rounded or triangular with 2 convex
sides and 3rd more flattened side
• flattened side has a depression in the middle
• 40s has a convex and flat side-partition on flat
side, it divides the smaller subunit into 2 equal
portion
• Smaller subunit is attached to larger one by
concave side
• Assumed that polypeptide chain, newly
synthesized in ribosome passes through the
channel of larger subunit into the cavity of ER

38. 70s structure

39. • In E.coli,50s subunit is bilaterally symmetrical in
front view ,shows 3 protuberance arising from
central rounded base
• Central one is prominent
• Transverse cleft divided 30s into 2 parts,smller
head and larger body
• This gives it the appearance of telephone receiver
• 50s is compared to arm chair.
• Long axis of 30s is oriented transversely to central
protuberance of 50s
• Tunnel is formed between the hollow of smaller
and seat of larger subunit

40. PROPERTIES 70s 80s
Occurrence Prokaryote Eukaryote
Sedimentation
coefficient
70s 80s
Size Smaller(20-30nm) Larger(25-35nm)
Molecular weight 3 million 4.5 million
Subunit Smaller 30s &
larger 50s
Smaller 40s &
larger 60s
RNA 3 types 4 types
Protein 50-60 molecule 70-80 molecule
RNA-protein Ratio 2:1 1:1
1st critical Mg2+
level
0.5 micromole 0.3-0.1 micromole
2nd critical Mg2+
level
0.3 micromole 0.15 micromole

42. Chemical composition
• Made up of RNA and protein
• 70s has 60% RNA and 40% protein
• 80s has 45% RNA and 55% protein
• No lipid
• Certain metallic ions are present

43. Ribosomal RNA
• 70s has 3 types-23s rRNA,16s rRNA and 23s and 55 rRNA
• 80s has 4 types-28s rRNA,5s and 5.8s rRNA and 18s rRNA
• 55s ribosome of mammalian mitochondria contain 21s and
12s rRNA

44. Ribosomal Protein
• Protein content is complex
• 50 to 55 proteins are isolated from ribosomal
subunit
• Some are called core protein-bind to rRNA
• Secondary binding protein-divided into acidic and
basic types
• Split proteins are released
• Many ribosomal protein act as enzymes and
catalyze protein synthesis

45. • They include :-
1. Initiation protein
2. Transfer protein
3. Peptide transferase
4. Termination factor
5. Metallic ions