Basic description of these organelles, their characteristic features, their functions and with special emphasis on their protein targeting

1. PRESENTER : J. AISHWARYA
M.Sc. 3RD YEAR MEDICAL BIOCHEMISTRY
18-May-21 1

2. MITOCHONDRIA
18-May-21 2

3. MITOCHONDRION - HISTORY
• A double membrane bound organelle found in cytosol of eukaryotic cells
• Mito – thread, chondrion – granule like
• First observed - Richard Altman ( 1894)
• Term mitochondria was coined by Carl Benda (1898)
MAIN FUNCTION:
• They produce enzymes for the metabolic conversion of
food to energy.
18-May-21 3

4. ORIGIN OF MITOCHONDRIA
• Derived from bacteria – process by Endosymbiosis
• Mitochondria arose - 2 billion years ago when a bacterium fused with an archael
cell / established a symbiotic relationship with a primitive eukaryotic cell
• The closest extant relatives of Bacteria that gave rise to mitochondria - Rickettsia.
• The first person to recognize mitochondria as descendents of endosymbiotic
bacteria - Ivan
18-May-21 4

5. THE ENDOSYMBIOTIC THEORY
18-May-21 5

6. EVIDENCES TO SUPPORT ENDOSYMBIOTIC THEORY
• Mitochondria – self replicating bodies like bacteria, divide by binary fission
• Two membranes , inner membrane composition similar to bacteria
• Mitochondrial DNA – similar to bacterial DNA
• mi ribosomes, enzymes & transport proteins – similar to bacteria
• similar size
• Antibiotics – inhibits mitochondrial protein synthesis
18-May-21 6

7. MORPHOLOGY
• Size 0.05 – 1.0 µm in diameter
• Length 1 – 10 µm long
• Shape Bean shaped , in fibroblast it is
elongated and thread like.
• Number Depends on type, size and
functional state of cell.
Eg : an average liver cell contain around
1500 mitochondria.
• Location Cells with high energy
requirement
18-May-21 7

8. GENERAL PROPERTIES
• Mitochondria occupy - 20% of the cytoplasmic volume of a eukaryotic cell.
• Often depicted as short, bacterium-like bodies - diameter of 0.5–1 μm
• Remarkably dynamic and plastic, moving about the cell, constantly changing
shape, dividing, and fusing
• Mitochondria are often associated with the microtubular cytoskeleton - which
determines their orientation and distribution in different cell types.
18-May-21 8

9. • In highly polarized cells - neurons, mitochondria can move long distances (up
to a meter or more ) being propelled along the tracks of the microtubular
cytoskeleton.
• In other cells, mitochondria remain fixed at points of high energy demand- for
example, in skeletal or cardiac muscle cells, they pack between myofibrils, and
in sperm cells they wrap tightly around the flagellum
18-May-21 9

10. SHAPE, DISTRIBUTION
• In higher plants :
• Rod shape with hemispheroidal ends, Some
are cup or filamentous shape.
• Vary from globular to threadlike or branched
• In Animals
• Long filaments , not spatial in arrangement
18-May-21 10

11. NUMBERS
• Depends on what the cell needs to do
• Flagellated protozoa or sperm, they are found around the base of the flagellum.
• Cardiac muscle, they surround the contractile parts.
• Hummingbird flight muscle is the richest sources of mitochondria.
When energy is not enough, more mitochondria are created ,they grow, move,
and combine with other mitochondria
18-May-21 11

12. INTERACTION WITH OTHER MEMBRANE
• Contacts between mitochondria and ER - facilitate the
exchange of lipids between the two membrane systems.
• Facilitate fission and fusion - involved in the distribution
and partitioning of mitochondria within cells
18-May-21 12

13. STRUCTURE
18-May-21 13

14. • Double membrane
• creates 3 compartments –
• OUTER MEMBRANE
• INTERMEMBRANE SPACE
• INNER MEMBRANE
• CRISTAE
• MATRIX
• The outer and inner membrane is composed of phospholipid bilayers and
proteins.
• The two membranes have different properties.
18-May-21 14

15. OUTER MEMBRANE
• Simple phospholipid bilayer.
• Contain large number of integral protein structures called porins, which allows
molecules to freely diffuse from one side of the membrane to the other.
• Porins pass molecules less then 5000 D - a special class of β-barrel-type
membrane protein that creates aqueous pores across the membrane
• Ions, nutrient molecules, ATP, ADP etc can pass through the outer membrane
with ease.
• The outer mitochondrial membrane is composed - 50% phospholipids by
weight and contains a variety of enzymes
18-May-21 15

16. INNER MEMBRANE SPACE
• It is also known as Perimitochondrial space.
• It has high proton concentration.
• The space is approximately 70 A.
• Because the outer membrane is freely permeable
to small molecules- concentration of small
molecules such as ions and sugars in the
intermembrane space is same as that of the
cytosol.
• Proteins present, participate in ATP synthesis
18-May-21 16

17. INNER MEMBRANE
• Is freely permeable only to oxygen, CO₂ , H₂O.
• The inner mitochondrial membrane contains proteins that perform redox
reactions in oxidative phosphorylation, ATP synthase, transport proteins,
protein import machinery, mitochondria fusion and fission protein.
• Several antiport systems exist , allowing exchange of anions between the
cytosol and the mitochondrial
18-May-21 17

18. CRISTAE
• Are folds of inner mitochondrial membrane, which expand its surface area ,
enhancing its ability to produce ATP.
• Stalked particles or inner membrane spheres : cristae is covered with this inner
membrane spheres called stalked particles or knobs or heads.
18-May-21 18

19. MATRIX
• It is the space enclosed by the inner membrane.
• Gel like consistency ,Dense , homogenous.
• Contains 2/3 rd of total protein of mitochondria.
• Matrix have enzymes, DNA genome, ribosomes, tRNA, granules, fibrils and
tubules.
• The matrix is important in the production of ATP with the aid of the ATP
synthase contained in the inner membane.
• Major enzymes include enzymes involved in - Synthesis of nucleic acid and
proteins, Fatty acid oxidation.
18-May-21 19

20. MITOCHINDRIAL DNA (mt DNA)
• Small, Double stranded ,covalently closed ,circular molecule.
• Occurs in multiple copies
• It has 16569 bp , 37 genes
• Most usually remains attached to inner mitochondrial membrane.
• Stores biological info required for growth and multiplication of mitochondria.
• Encode RNA s and proteins - essential for mitochondrial function.
• It codes 2rRNAs , 22 tRNAs and 13 mitochondrial membrane proteins.
• Can undergo replication and duplication
18-May-21 20

21. 18-May-21 21

22. BIOGENESIS
• They grow - importing most of their proteins from the cytoplasm and by internal
synthesis of some proteins and replication of the genome.
• Similar to cells, mitochondria divide and fuse with other mitochondria
maintaining their number of cells.
• The balance between fusion and fission - major determinant of mitochondrial
number, length, and degree of interconnection
• Fusion > fission, the mitochondria tend to become more elongated and
interconnected
• Fission > fusion ,- more numerous and distinct mitochondria.
18-May-21 22

23. TARGETING OF MITOCHONDRIAL PROTEINS
18-May-21 23

24. • Mitochondrial proteins synthesized – 80S cytosolic & 70S matrix ribosomes
• 99 % of mit proteins – synthesized as precursors in cytosol
• translocated post-translationally
• Mit protein import – membrane receptos & translocons
• initiated – binding of mit targeting sequence to import receptor in outer
membrane
• TOM (translocase of outer membrane) & TIM (translocase of inner
membrane)
• Cooperative mechanism
TARGETING OF MITOCHONDRIAL PROTEINS
18-May-21 24

25. 18-May-21 25

26. TOM COMPLEX
TIM COMPLEX
18-May-21 26

27. Mitochondrial targeting signal sequences
• Signal sequences – 2 classes
• N-terminal cleavable matrix targeting sequence
• Non- cleavable internal targeting sequence
• Presequences – rich in basic & hydroxyl AA, 10-30 AA residues
18-May-21 27

28. 18-May-21 28

29. 1. Targeting mit proteins to matrix
• Matrix proteins contain presequences
• synthesized by 80S cytosolic ribosome
• maintained in unfolded state – cytosolic chaperone Hsp70
18-May-21 29

30. 18-May-21 30

31. 2. Targeting mit proteins to IMM
• similar to targeting proteins to matrix side – except there is an additional stop
transfer signal – located after presequence
• arrests translocation
• OXA complex – insert proteins synthesized by 70S ribosome
18-May-21 31

32. 3. Targeting to outer membrane
18-May-21 32

33. MITOCHONDRIAL
DISORDERS
18-May-21 33

34. 18-May-21 34

35. DIAGNOSIS
18-May-21 35
• Unfortunately, mitochondrial genetic disorders can be difficult to diagnose, and many affected
people may never receive a specific diagnosis.
• In some cases, the pattern of symptoms may be suggestive of a specific mitochondrial
condition. If the disease-causing gene(s) associated with the particular condition is known, the
diagnosis can then be confirmed with genetic testing.
• In these cases, a physician may evaluate the levels of certain substances in a sample of blood
or cerebrospinal fluid.
• Exercise testing
•Magnetic resonance spectroscopy (detects abnormalities in the brain's chemical makeup)
•Imaging studies of the brain such as MRI or CT scan
•Electroencephalography (EEG)
•Tests that evaluate the heart including electrocardiography and echocardiography
•Muscle biopsy

36. 18-May-21 36

37. PEROXISOME
History of Peroxisomes
• First observed by electron microscopy in animal cells (1950s), then in plant cells
(1960s)
• Christian deDuve (1965) - Isolated from liver cells by centrifugation
• Called them peroxisomes because they generate and destroy H2O2
18-May-21 37

38. ORIGIN
• Peroxisomes are a vestige of an
ancient organelle that performed
all the oxygen metabolism in the
primitive ancestors of eukaryotic
cells.
• When the oxygen produced by
photosynthetic bacteria first
accumulated in the atmosphere, it
would have been highly toxic to
most cells.
• Peroxisomes might have lowered
the intracellular concentration of
oxygen.
18-May-21 38

39. • Single membrane
• Roughly spherical
• 0.2 - 1.7µm
• Composition varies
• Peroxisomes are also called Microbodies
• They also resemble lysosomes in being filled with enzymes
• They are self replicating
• contain enzymes to oxidize organic substances like fats
• Hydrogen peroxide is broken down right away by the enzyme catalase into
oxygen and water.
• Peroxisomes are abundant in the liver where they produce bile salts and
cholesterol and break down fats
PROPERTIES
18-May-21 39

40. • do not contain DNA or ribosomes
• all of their proteins are encoded in the nucleus
• They contain oxidative enzymes, such as catalase and urate oxidase, at such
high concentrations that the peroxisomes stand out in electron micrographs
because of the presence of a crystalloid protein core
18-May-21 40

41. 18-May-21 41

42. 18-May-21 42

43. Peroxisomes Use Molecular Oxygen and Hydrogen
Peroxide to Perform Oxidation Reactions
• RH2 + O2 → R + H2O2
• removes H2 atoms
• H2O2 + R′H2 → R′ + 2H2O.
• This type of oxidation reaction important in liver and kidney cells, where
the peroxisomes detoxify various harmful molecules that enter the
bloodstream.
• In addition, when excess H2O2 accumulates in the cell, catalase converts it
to H2O through the reaction 2H2O2 → 2H2O + O2
18-May-21 43

44. FUNCTIONS
• Fatty acid oxidation
• Bile acid synthesis
• lipid biosynthesis – cholesterol & dolichol
• plants – two pathways
18-May-21 44

45. GLYOXYLATE
CYLCE
PHOTORESPIRATION
18-May-21 45

46. PROTEIN TARGETING IN PEROXISOME
• A Short Signal Sequence Directs the Import of Proteins into Peroxisomes
• A specific sequence of three amino acids (Ser–Lys–Leu) located at the
C-terminus of peroxisomal proteins functions as - import signal
• transport from cytosol to peroxisome – post translationally
• proteins synthesized in membrane free ribosomes
• Peroxins – 23 types
• ATP dependent process
• Sequence- Ser – Lys - Leu
18-May-21 46

47. 18-May-21 47

48. 18-May-21 48

49. 18-May-21 49

50. NUCLEUS
18-May-21 50

51. NUCLEUS
• light microscope - nucleus is the largest visible compartment
• The presence of a nucleus distinguishes eukaryotic cells from prokaryotic cells
• Houses all of the eukaryotic cell’s genome and acts as a center for controlling
cellular activities
18-May-21 51

52. STRUCTURE
• A double membrane called nuclear envelope encloses the nucleus.
• The lumen separates the two membranes and is continuous with the
Endoplasmic Reticulum.
• Macromolecules pass between the nucleus and cytoplasm through the Nuclear
Pore complexes (NPCs) - channels spanning the envelope.
• Nucleolus is the most clearly visible structure.
• Regions other than the nucleolus are referred to as the nucleoplasm.
• Other sub-compartments include speckles, cajal bodies and PML bodies etc.
• Inside the nucleus the DNA can be found in the compacted and highly stained
form, heterochromatin or in the less densely compacted form the euchromatin.
18-May-21 52

53. NUCLEAR MEMBRANE
• Nuclear Membrane is a fence between nucleus and cytoplasm to stave off free
transmission of molecules.
• Provides nucleus an identity of separate biochemical compounds.
• The nuclear membrane consists of:
• 1. Outer nuclear membrane
• 2. Inner nuclear membrane
• 3. Perinuclear space
• 4. Nuclear pores
• 5. Nuclear lamina
18-May-21 53

54. OUTER NUCLEAR MEMBRANE
• The outer nuclear membrane is continuous with endoplasmic reticulum,
therefore the lumen of nuclear membrane is directly connected with lumen of
ER.
• The outer nuclear membrane is functionally homologous to ER membrane.
• The cytoplasmic surface of outer nuclear membrane has ribosomes that are
different in composition of protein and these ribosomes are enriched in
membrane proteins (for cytoskeleton binding)
18-May-21 54

55. PERINUCLEAR SPACE
• Space is present between ONM and INM and is called Perinuclear space or
lumen of envelope.
• The thickness of each nuclear membrane is 7-8nm thick while perinuclear
space is 20-40nm thick.
18-May-21 55

56. INNER NUCLEAR MEMBRANE
• Proteins that are specific to nucleus are present in INM such as those that bind
the nuclear lamina.
• Including Lamin B receptor (LBR), lamin aassociated polypeptide (LAP) 1, LAP2,
emerin, MAN1 and nurim.
• Most of these proteins interact with lamins and chromatin.
• Mutations in emerin and nuclear lamins have been associated with muscular
dystrophies and lipodystrophy.
• Integral proteins of the inner nuclear membrane are synthesized on the rough
ER and reach the inner nuclear membrane by lateral diffusion in the connected
ER and nuclear envelope membranes.
18-May-21 56

57. WHAT ARE NUCLEAR PORES?
• Small polar molecules, ions and macro-molecules can only move in between the nucleus
and cytoplasm through channels.
• The large circles with diameter of 120nm and molecular mass of ̴125 million Dalton and
30X the size of ribosomes are pores that are collectively called as Nuclear Pore complex.
• They are composed of several proteins of 30 different types.
• Those specialized proteins are named as Nucleoporins
• The complex consists of an assembly of eight spokes attached to rings on the cytoplasmic
and nuclear sides of the nuclear envelope.
• The spoke-ring assembly surrounds a central channel containing the central transporter.
• Cytoplasmic filaments extend from the cytoplasmic ring, and filaments forming the
nuclear basket extend from the nuclear ring.
18-May-21 57

58. NUCLEAR PORE CHANNEL (NPC)
• The Phospholipid bilayer is only
permeable for non-polar
micromolecules.
• The only channel through which
transmission of polar micromolecules
and macromolecules occurs is through
Nuclear Pore Complex.
• NPCS are the points where lNM and
ONM are continuous
18-May-21 58

59. NUCLEAR LAMINA
• In multicellular eukaryotes, a fibrous mesh work supports the inner nuclear
membrane called Nuclear Lamina
• The nuclear lamina is present inside the nuclear envelope.
• Lamins are 60-80 kilo Dalton fibrous proteins that makeup the nuclear lamina
• Some associated proteins are also present.
• Lamins belong to a class of intermediate filament proteins.
• Nuclear Lamina disease:
• 1. Emery-Dreifuss muscular dystrophy
• 2. Hutchinson-Gilford progeria syndrome
18-May-21 59

60. TRANSPORT THROUGH NUCLEAR PORE COMPLEX
• Proteins < 50kDa – passively pass
• Proteins > 50kDa – actively transported
• proteins – imported & exported – signal sequence - Nucleus Localisation
signal (NES / NIS )
• Nuclear import receptor – Importin
• Nuclear export receptor – Exportin
• transported post-translationally & in fully folded confirmation
• NES - Leu rich
• NIS – Lys rich
• Karyopherins – homologous sequences
• Interact with FG (phe, gly) repeats of nucleophorins
18-May-21 60

61. 18-May-21 61

62. DISORDERS
Cancer associated morphological
changes:
• The most commonly used
quantitative nuclear
morphometric parameter is the
measurement of nuclei size.
• The nuclear-cytoplasmic ratio
(also known as the
“karyoplasmic ratio”)
18-May-21 62

63. LAMINOPATHIES
• group of diseases caused by alterations in
nuclear morphology are laminopathies
• genetic disorders arising from mutations in
the genes encoding lamins or lamin-
interacting proteins, such as NE integral
membrane proteins, and components of the
LINC complex
18-May-21 63

64. 18-May-21 64

65. 18-May-21 65

66. 18-May-21 66

67. 18-May-21 67

68. 18-May-21 68

69. 18-May-21 69