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transcriptron.pptx - Slide 1

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    • 1. RNA synthesis:transcrition • Transcription is the process of the synthesis RNA molecule.
    • 2. Transcription unit • Promoter region • Transcribed region • And termination region.
    • 3. Promoter region • Which is certain sequence of base located at the beginning of stretch. It is important for initiation for transcription to occur and can be recognized by RNA polymerase.
    • 4. prokaryotes • 1. The pribno box :stretch of 6 nucleotides (TATAAT) located about 10 bases to the left of the transcription. Initiation site. • 2. A second nucleotide stretch (TGTTGACA) that located about 35 bases to the left of the transcription initiation site. • 3. 19 bases (nucleotide) in between two streches.
    • 5. Cont. • Both pribno box and TGTTGACA are region can be recognized by RNA polymerase.
    • 6. Eukaryotic promotor • 1. The Hoges or TATA box: stretch of nucleotide that is almost similar to prokaryotes,located almost 25 nucleotide to the left of the trancrition initiation site. • 2. CAAT box : located about 70 to 80 nucleotides to the left of the trasncription initiation site. • 3. 40 bases between the two stretches.
    • 7. Cont. • Transcription region: which is stretch of DNA that is to be transcribed into RNA molecules. • Termination region: which is a stretch of DNA located at the end of DNA sequence to be trasncribed.
    • 8. Requirement for RNA synthesis • Transcription unit: transcription occurs on one of the two strands of the DNA template and never on both complementary strand • Four ribonucleotide triphosphate;ATP,GTP,UTP and CTP. • RNA poymerase ,RNA(=DNA –dependent RNA polymerase)
    • 9. Prokaryotes RNA polymerase. • 1.E.g E.coli ,enzyme synthesize all types of RNA molecules on DNA template. • 2. the holoenzyme of the RNA polymerase consist of a core molecule and specific protein factor(sigma [σ] factor. • A. The core enzyme molecule : 4 subunits: 2 of them are identical (alpha subunits.the other 2(β and β’) are similar but not identical .RNAP consist of 2 zinc molecules.
    • 10. Sigma factor • Enables the polymerase to recognize promotor regions on DNA . • Helps the core enzyme to attach more tightly to the promotor site. • Some regions on DNA that signal the termination of transcription are recognized by the RNA polymerase itself. Others are recognized by specific termination factor e.g rho factor of E.coli.
    • 11. Steps RNA synthesis • Initiation • Elongation • termination
    • 12. Initiation • 1. RNA polymerase holoenzyme binds with the promotor area: • A . Sigma subunit ,enables polymerase polymerase to recognize promotor region on DNA. • B. Β̒subuint to the template. • C. B subunit binds to the nucleotide substrate.
    • 13. Cont. • 2. binding of RNA poymearse DNA template leads to local separation (unwinding)of the DNA double helix into sense and antisense strands. • 3. first nucleotide of RNA transcript at the initiation site is almost always purine.
    • 14. Elongations • 1.At the sense formation of RNA molecule begins at the 5̒end of core enzyme with release of sigma factor. • 2. Then elongation of the RNA molecule occurs from 5 ̒ to 3̒ end ,antiparallel to its template. • 3. nucleotide building blocks are 5̒ ribonucleotide triphosphate(ATP ,GTP,CTP,and UTP). They are inserted in the RNA = pairing rule.pyrophosphate is released when each new nucleotide is added to growing chain . GTPGMP + PPi
    • 15. Cont. • 4.RNA polymerase forms a phosphodiester bond between the ‘3” OH of one ribose sugar and 5’ OH of the next ribose. • 5. the process of elongation of RNA chain continues until a termination region reached.
    • 16. Termination: • Termination region on DNA template can be recognized by : • A. RNA – polymerase enzyme itself (rho independent termination),. • B. Rho [ρ] factor which may be required for the release of both RNA strand and RNA polymerase (rho dependant termination).
    • 17. Termination result from either • A. Binding of rho factor polymerase enzyme : when RNA – polymerase enzyme reaches the termination site,rho factor binds with it causing termination. • Slowing down of RNA – polymerase at the termination site: • 1. termination site on DNA is charaterized by the presence of palidromes.
    • 18. Cont. • Palidromes is a region of a double stranded DNA in which each of the two strands haas the same sequence when read in the same direction. E.g in the 5’ to 3’direction. • 2.The RNA transcript of the DNA palidrome can form a stable hairpin structure which is self –complementary structure. This hairpin structure causes slowing of RNA – poylmerase at the termination site.
    • 19. antibiotics • Rifampcin: binds to the beta subunit . Treatment of leprosy and TB patients. • Actinomycin D: binds to DNA template ,prevent movement of RNA – polymerase along the DNA. • Alpha –amanitin: toxin from mushroom , inactivates RNA – polymerase II of eukaryotes.
    • 20. Post transcriptional modification of RNA • A. eukaryotes mRNA : primary mRNAs are called heterogeneous nuclear RNA (hnRNA) .hnRNA is modified into mature mRNA in the nucleus by capping additon of nucleotides and splicing.
    • 21. 5’capping • The 5’end of the RNA requires a cap which is 7 methyl-guanosine triphosphate. It is attached by a 5’to 5’triphosphate linkage. • A. reaction needs an enzyme called guanyl transferase. • B. function of this cap is to facilitate the initiation of translation, and protects the 5’end of mRNA from attack by 5’to 3’exonucleases.
    • 22. Addition of poly (A) tail. • mRNA require almost 40 to 200 adenine nucleotides added at 3’terminus to form a poly adenine (A) tail: • poly A polymerase . Enzyme for the reaction. • This tail protects the 3’end of mRNA form 3’to 5’ exonuclease attack.
    • 23. Splicing (removal of introns) • hnRNA is formed many pieces of some of them (exons) will be translated into amino acids. • Others (introns) will be not be translated into amino acids and must be removed before translation takes place.
    • 24. spliceosomes • Responsible for removal of introns from the hnRNA , and splicing (ligation) of both ends of exons to form mature mRNA. • It consist of primary hnRNA ,4 small nuclear RNA’s (U1,U2,U5, and U4/U6) and an undetermined number of proteins. • Smaller nuclear RNA’s (snRNA) is to bind each end of the introns by forming base pair with each other.
    • 25. cont. • Spliceosomes facilitate also the transport of mature mRNA from the nucleus to the cytoplasm. • Defect in the process of spilicing may lead to disease e.g at least one from of beta thalassemia , a disease in which there is absent synthesis of beta chain of hemoglobin appears to result from a nucleotide change at an exon-introns junction.
    • 26. Transfer RNA(t RNA) • 1.It serves as a adapter mol. For the translation of mRNA into protein sequence. • 2. primary tRNA transcript are subjected to many modifications: • A. tRNA mol are transcribed as larger precusors. These precursors are reduced in size by specific class of ribonucleases. • B. Attachment of the characteristics C.C.A terminus at the 3’end of the molecules.
    • 27. Cont. • The tRNAs contain many modification of the standard bases A,U,G and C .Some bases are methylated, alkylated, or attached to carbohydrate residue by glucosidic bonds. • some tRNA contain near to the anticodon loop a single intron 10-40 nucleotides long. These introns are removed with splicing ofr exons to produce an active tRNA for protein synthesis.
    • 28. Ribosomal RNA(rRNA) • 1.In mammals cells ;rRNA is transcribed as a single larger precursor molecules called 45s. • 2.In the nucleus ,45s is methylated and cleaved by specific endonucleases and exonucleases to give four kind of r RNA : 5SrRNA,5.8SrRNA,18SrRNA and 28SrRNA.
    • 29. Cont. • 3. The four kind rRNA combine with a number proteins to form a ribosomes. • 1. ribosomes are cytoplasmic nucleoproteins composed of 4 rRNAs plus a number of proteins. • 2. These rRNAs and proteins are distributed specifically between the two smaller and larger ribosomal subuints.
    • 30. Cont. • 3. The smaller subunits is called 40S .It contains one 18S rRNA and 33 proteins. • 4. The larger subunit is called 60S . It contains the remaining r RNAs (28S,5.8S and 5S) and 45 proteins. • 5. Both smaller (40S) subuint form the whole 80S ribosome.
    • 31. Function of ribosomes • They are site of protein synthesis within cells.