MITOCHONDRIA AND RIBOSOME

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

8. 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

9. 2 conformational stage for mitochondria : Orthodox stage -ADP low, matrix more, cristae well defines and no phosphorylation 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

10. 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

11. 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.

12. 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

13. 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

14. 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 1 st 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

15. 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 1 st step in the common pathway for degradation of fuel molecules It undergoes 2 transformation stage

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

17. 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

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

19. Mitochondrial DNA

20. 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 .

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

22. 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

23. 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

24. Functions Respiration or oxidation of food stuffs Transduction of energy Synthesis of protein Yolk formation ATP transport Formation of mitochondrial spiral Lipid synthesis Extra chromosomal inheritance Heat production Storage

25. RIBOSOME

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

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

28. 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

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

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

31. 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

32. 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

33. Ultrastructure-80s

34. 60s appear rounded or triangular with 2 convex sides and 3 rd 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

35. 70s structure

36. 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

37. 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 1 st critical Mg2+ level 0.5 micromole 0.3-0.1 micromole 2nd critical Mg2+ level 0.3 micromole 0.15 micromole

38. 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

39. 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

40. 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

41. They include :- Initiation protein Transfer protein Peptide transferase Termination factor Metallic ions