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Dna rna-13

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  • 1. DNA structure determination10/10/0533• Franklin collected x-raydiffraction data (early 1950s)that indicated 2 periodicitiesfor DNA: 3.4 Å and 34 Å.• Watson and Crick proposed a3-D model accounting for thedata.
  • 2. CELL STRUCTURE ANDBIOMOLECULESMolecular biology- Energy based flow ofinformation from DNA to RNA toproteins and enzymes i.e. from Hypopituitary access to hormones tohomeostatic etc.
  • 3. The mother of all biomolecules10/10/05421ffkLarge subunit of the ribosome(proteins at least)
  • 4. Chromatin Structure
  • 5. INACTIVE &ACTIVE CHROMATIN• Active (transcriptionally active)differfrom from inactive region.• DNA in active region contain largeregions(100,000bp)prone to digestion byDnase- I (single strand cut)
  • 6. MAJOR AND MINORGROOVES• MINOR– EXPOSES EDGE FROM WHICH C1’ ATOMSEXTEND• MAJOR– EXPOSES OPPOSITE EDGE OF BASE PAIR• THE PATTERN OF H-BOND POSSIBILITIES ISMORE SPECIFIC AND MOREDISCRIMINATING IN THE MAJOR GROOVE
  • 7. DNA structure10/10/0534Fig. 8-15•DNA consists of twohelical chains woundaround the same axis ina right-handed fashionaligned in an antiparallelfashion.• There are 10.5 base pairs,or 36 Å, per turn of thehelix.• Alternating deoxyribose andphosphate groups on thebackbone form the outsideof the helix.• The planar purine andpyrimidine bases of bothstrands are stacked insidethe helix.
  • 8. Much of Genome is notTRANSCRIBED• The entire human haploidgenome contain sufficent DNA tocode for app. 1.5 million genes.• Human genome encodes lessthan 100,000 proteins ie 1% ofhuman genome.• 24% of genome as Introns
  • 9. More than half DNA of Eukaryotic isnon Repetitive Sequences.• In humans 10,000 to 15000 . genes areexpressed.• Different combination of genes areexpressed in each tissue,of course andhow this is accomplished in one of themajor unanswered question in biology.
  • 10. 30% of Human Genome hasRepetitive sequences• A.Highly repetitive: 5-500 bp lengthrepeated many times.(these seq. aretranscriptionaly inactive)may play role instructure of ch.• B. Moderately repetitive: 106copiesper haploid genome are not clustered butare interspred with unique seq.• C. Microsatelite repeat seqs.2-6bprepeated up to 50 times(AC=TG)ACrepeat seqs. Are50000-100000 locationsin genome.
  • 11. TELOMERE• The ends of each chromosomecontains structures called TELOMERES• Telomere consist of short repeat TGsequences.(5’-TTAGGG-3’)can run forkb.• TELOMERASE ( RNA+RNA dependentDNA polymerasesor{ Reversetranscriptase} is responsible fortelomere synthesis) & maintains lengthof telomere.
  • 12. Cont………………………….• Genome of Prokaryotes arecircular.human &Eukaryotes is linear asresult the lagging strand has incomplete51 end. Each round of replication wouldshorten the chromosome.Telo=End.• How are repeated sequences generatedthe enzyme TELOMERASE performs thisfunction.• The protein component oftelomerases,hence it acts as reversetranscriptase.that carries its own template.• Telomerase levels are raised in cancercells.
  • 13. DNA• Two helical polynucleotide chainsare coiled around a common axis.• The chains run in oppositedirections.(Template and Coding)• Sugar-Phosphate backbones areon the outer side and Purine&Pyrimidene bases lie on theinside of helix.• The bases are prepandicular to thehelix axis. (10 bases per turn ofHelix)• Diameter of helix is 20 Ao.
  • 14. Cont………………………..• In between large regions thereare shorter streches (100-300)which are more sensativeto Dnase-I These sites providesa sight for transcription.(euchromatin)• Transcriptionally inactive ch. Isdensly packed.(Hetrochromatin)
  • 15. A-DNA• RIGHT-HANDED HELIX• WIDER AND FLATTER THAN B-DNA• 11.6 BP PER TURN• PITCH OF 34 A  AN AXIAL HOLE• BASE PLANES ARE TILTED 20 DEGREESWITH RESPECT TO HELICAL AXIS– HELIX AXIS PASSES “ABOVE” MAJOR GROOVE  DEEP MAJOR AND SHALLOW MINORGROOVE• OBSERVED UNDER DEHYDRATINGCONDITIONS
  • 16. B-DNA• B-DNA Right handed double helix.• Supercoils store energy.• Supercoiled DNA is prefered form.• DNA of E.coli is1.4mm, 4.7millionbp.• Eukaryotes contain more than tentimes of DNA as compaired toprok.
  • 17. Z-DNA• A LEFT-HANDED HELIX• SEEN IN CONDITIONS OF HIGH SALTCONCENTRATIONS– REDUCES REPULSIONS BETWEEN CLOSESTPHOSPHATE GROUPS ON OPPOSITESTRANDS (8 A VS 12 A IN B-DNA)• IN COMPLEMENTARY POLYNUCLEOTIDES WITHALTERNATING PURINES AND PYRIMIDINES– POLY d(GC) · POLY d(GC)– POLY d(AC) ⋅ POLY d(GT)• MIGHT ALSO BE SEEN IN DNA SEGMENTS WITHABOVE CHARACTERISTICS
  • 18. B,A and Z DNA10/10/0538Fig. 8-19• B form - The most commonconformation for DNA.• A form - common for RNAbecause of different sugarpucker. Deeper minor groove,shallow major groove• A form is favored in conditionsof low water.• Z form - narrow, deep minorgroove. Major groove hardlyexistent. Can form for someDNA sequences; requiresalternating syn and anti baseconfigurations.36 base pairsBackbone - blue;Bases- gray
  • 19. DNA strands10/10/0537Fig. 8-16• The antiparallel strands of DNAare not identical, but arecomplementary.• This means that they arepositioned to align complementarybase pairs: C with G, and A withT.• So you can predict the sequenceof one strand given the sequenceof its complement.• Useful for information storageand transfer!• Note sequence conventionally isgiven from the 5 to 3 end
  • 20. Discovering the structure of DNA• DNA = Deoxyribose nucleic acid• Made out of sugars (deoxyribose), phosphatesand nitrogen bases
  • 21. Difference betweenDNA and RNA• Sugar: DNA;deoxyribose.• RNA; Ribose.• DNA: A,T,G,C.: RNA;A,U,G,C.• DNA Double,RNA single stranded.• DNA A=T,G=C, RNA; U=A,,G=C not.• RNA can be hydrolysed byAlkali,DNA not.
  • 22. HISTONES:• H1• H2A• H2B• H3• H4Acetylation linked with replication,H1with condensation,DNArepair,transcriptionrepression,methylation of histonesactivation and repression of gene.
  • 23. HISTONES• Most abundant basic chr.protein.• H1Loosly bound to chr,&not notnecessary,but adjecent nucleosomeis joined by H1, nucleosome• Nucleosome contain four types ofhistones.H2A,H2B,H3 and H4• Function identical in all eukaryotes.• Four core Histones are subject to 6types of covalent modifications
  • 24. HISTONES• Acetylation,Methylation,Phosphorylation,ADP-ribosylation,monoubiquitylationand Sumoylation.• H3,H4 form tetramer• h2A &H2B form Dimer.Underphysiologic conditionsolig.associate & form octamer.• The amino tail(His) is available co-valent modifications• One octamer has 1.75 turns of DNA(146bp)
  • 25. The mother of all biomolecules10/10/05421ffkLarge subunit of the ribosome(proteins at least)
  • 26. Z-DNA• 12 (W-C) BASE PAIRS PER TURN• A PITCH OF 44 DEGREES• A DEEP MINOR GROOVE• NO DISCERNIBLE MAJOR GROOVE• REVERSIBLE CHANGE FROM B-DNA TOZ-DNA IN LOCALIZED REGIONS MAY ACTAS A “SWITCH” TO REGULATE GENEEXPRESSION– ? TRANSIENT FORMATION BEHINDACTIVELY TRAN-SCRIBING RNA POLYMERASE
  • 27. FORCES THAT STABILIZENUCLEIC ACID STRUCTURES• SUGAR-PHOSPHATE CHAINCONFORMATIONS• BASE PAIRING• BASE-STACKING,HYDROPHOBIC• IONIC INTERACTIONS
  • 28. DNA TOPOLOGY• THE TOPOLOGICAL PROPERTIES OF DNA HELP USTO EXPLAIN• DNA COMPACTING IN THE NUCLEUS• UNWINDING OF DNA AT THE REPLICATION FORK• FORMATION AND MAINTENANCE OF THETRANSCRIPTION BUBBLE• MANAGING THE SUPERCOILING IN THEADVANCING TRANSCRIPTION BUBBLE
  • 29. mRNA (messenger RNA)• mRNA is synthasized from DNA byenzyme RNA Polymerase.• Introns and Exons.(HetrogenoushnRNA)• Most hetrogenous,highly elongatedand short lived.• Has 5’—3’ polarity complimentary tocoding strand.• For protein synthesis.• Stable in Euk. Unstable in Pro.
  • 30. tRNA (Transfer RNA)• Much smaller in size (75-90ribonucleotide)Clover leaf shaped.• 15% of total cellular RNA.• 20 species of tRNA.• It acts as adaptor molecule.• 40 different tRNA.• Five arms of tRNA: 1:D-Arm; 2:TUC-Arm 3: CCA Arm(Acceptor) 4:Anticodon Arm 5: Variable Arm orExtra Arm (determine Sp.).
  • 31. tRNA• Amino Acid acceptor arm 7 base pairstem• 5’—3’ CCA.Links Amino Acid.AA arelinked co-valently to correspondingtRNA by enz.AAtRNA Synthetase.• Anticodon Arm Has 5-bp loop andtriplet nucleotidesequence(anticodon) complimentaryto codon.• DHU arm .3-4 base pairs.• TUC arm stem has 5 bp.
  • 32. RIBOSOMAL RNA• Are present in Ribosomes inassociation with many polypeptides.• 80 % of total cellular RNA.• Very complex in Euc. Two sub-units(60S)and (40S)• Protein Synthesis.• Larger unit has 3 RNA ie 5S,5.8S,28Swith 50 polypeptides.• Smaller subunit has 18S and 30polypeptides.
  • 33. MITOCHONDRIAL DNA• 54 out of 67 genes are coded bynuclear genes,rest are coded byMitochondria.(mt)• It form 1% of total cellular DNA.• It codes 13 proteins that play key rolein the respiratory chain.
  • 34. Features of mtDNA• Circular,double stranded• Contains 16,569 bp• Encodes 13 protein subunits.• Encodes 22 mt. tRNA molecules.• Encodes(16s) and a small (12s)mt rRNA• Very few untranslated sequences.• High mutation rate (5-10 times to nucl.)• M for M (Maternaly inherited Mitochondria)
  • 35. Discovering the structure of DNA• DNA = Deoxyribose nucleic acid• Made out of sugars (deoxyribose), phosphatesand nitrogen bases
  • 36. The mother of all biomolecules10/10/05421ffkLarge subunit of the ribosome(proteins at least)
  • 37. • MitochondriaFunction- ATP synthase, fatty acid oxidation, Krebs cycle, cytosolicCa2+regulation.Marker- Succinate dehydrogenase, glutamate dehydrogenase.
  • 38. Redox potential of some redox systemRedox system Eo`(volt)NADH/NAD+-0.32Succinate/Fumarate -0.32FADH2/FAD 0.0Coenzyme Q.H2/CoQ 0.10Cytochrome b (Fe2+/Fe 3+) 0.12Cytochrome c1 (Fe2+/Fe 3+) 0.22Cytochrome a (Fe2+/Fe 3+) 0.29Cytochrome a3 (Fe2+/Fe 3+) 0.39HO2/1/2O2 0.82
  • 39. • Endoplasm reticulum (2 type RER & SER)Function- Production of excretory proteins, glycosylation, drugmetabolism.Marker- Glucose-6-phosphatase.• Golgi complexFunction- Synthesis of lipoprotein, formation of lysosomes, secretionof proteins.Marker- Galactosyle transferase, Fucosyle transferase.• PeroxisomesFunction- Fatty acid oxidation, detoxification of H2O2.Marker- Catalase, uric acid oxidase.
  • 40. • LysosomesFunction- Intracellular digestion and detoxification.Marker- Acid phosphatase.