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TRANSLATION

TRANSLATION. Dr.S.Chakravarty MD. Site :-Ribosomes in Cytoplasm. THE CENTRAL DOGMA. TRANSLATION. Dr.S.Chakravarty , MBBS,M.D. Structure of t-RNA. Antiparallel to the coding sequence of amino acid on the m-RNA. Recognition of

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TRANSLATION

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  1. TRANSLATION Dr.S.Chakravarty MD

  2. Site :-Ribosomes in Cytoplasm

  3. THE CENTRAL DOGMA

  4. TRANSLATION Dr.S.Chakravarty , MBBS,M.D

  5. Structure of t-RNA Antiparallel to the coding sequence of amino acid on the m-RNA. Recognition of t-RNA to the enzyme aminoacyl t-RNA synthatase Binding of aminoacyl-tRNA to ribosomal surface at the site of protein synthesis. Site of attachment of specific amino acid Sequence of code in anticodon arm : 3’-variable base-modified purine-XYZ-pyr, pyr-5’

  6. tRNAs • 75–90 nucleotides long and contain posttranscriptionally modifiedbases. • 2-dimensional structure of tRNAs is a cloverleaf.

  7. Activation of t-RNA Step 1: amino acid + ATP → aminoacyl-AMP + Ppi Step 2: aminoacyl-AMP + tRNA → aminoacyl-tRNA + AMP Twenty different aminoacyl t-RNA synthases for twenty different aminoacids

  8. Translation of mRNA Can Be Divided into Three Steps • Initiation • Elongation • Termination

  9. Initiation requires: • the small and large ribosomal subunits • mRNA • GTP • charged initiator tRNA • initiation factors

  10. Protein synthesis – Eukaryotes • Initiation • Elongation • Termination • m-RNA is read from 5’ 3’ end. • m-RNA is identified by 18 s RNA through 7 methyl guanosine CAP

  11. Initiation • Formation of 43 s pre-initiation complex: a)First the ribosome dissociates into into 40S and 60S subunits. 40 S subunit 5 3 1A Initiation factors eIF3 and eIF1A and eIF5 bind to the dissociated 40 S unit GTP eiF2 Binary complex met tRNAi 40 S subunit 5 43 s pre-initiation complex 3 1A 2

  12. Role of eIF-2 • eIF2 consists ofα , β and ʏ subunits • eIF-2α is phosphorylated by several protein kinases. • Phosphorylation by at least four different protein kinases during stress: • Starvation • Viral infection • Abnormal protein (misfolded proteins) • Hyperosmolarity • Heat shock • Phosphorylated eIF-2α binds tightly to and inhibits eIF-2β which is required for regeneration of GTP from GDP on factor 2.This blocks protein synthesis. • Insulin increases protein synthesis by removing the phosphate and inactivating eIF-2a

  13. 2. Formation of 48s initiation complex 40 S subunit 5 CAP (A)n 3 4F =4E+ 4G4A ATP 1A 2 4F CAP (A)n 4B ATP Cap helps in binding m- RNA to 43S preinitiation complex 4F CAP (A)n M-RNA 4B PAB 4F (A)n CAP 5 PAB 3 1A 2 48s initiation complex

  14. Kozak Consensus Sequence is a sequence which occurs on eukaryoticmRNA and has the consensus (gcc)gccRccAUGG. The Kozak consensus sequence plays a major role in the initiation of the translation process

  15. Formation of 80s initiation complex 48s initiation complex 4F 2 Binary complex CAP 5 (A)n PAB 3 GTP 1A 2 2 2B 60 s unit GDP eIF 5 2 (-) AUG CAP (A)n PAB 2α A-site p Met Kozak consensus sequences P-site

  16. Peptide bond formation Enzyme-Peptidyl transferase

  17. termination HYDROLYSIS

  18. P bodies • Nontranslating mRNAs Can Form Ribonucleoprotein Particles that Accumulate in Cytoplasmic Organelles Termed P Bodies • These associate with the protein constituents that form P bodies, small dense compartments that incorporate mRNAs as mRNPs. • Over 35different proteins are present (RNA helicases, RNA exonucleasesetc) • Certain mRNA are temporarily stored and then retreived and utilized for translation.

  19. PROKARYOTES • Polysomes (or polyribosomes) are mRNAs with several ribosomes translating at once.

  20. Variation in Protein Structure Provides the Basis of Biological Diversity • Following translation, polypeptides fold up and assume higher order structures, and they may interact with other polypeptides.

  21. Antibiotic Sites of Action: RNA synthesis (RNA polymerase inhibitors) DNA replication (topoisomerase/gyrase inhibitors) Rifampin Fluoroquinolones Protein synthesis (30S inhibitors) Protein synthesis (50S inhibitors) Tetracycline Kanamycin Gentamicin Streptomycin Erythromycin Chloramphenical Clindamycin Fusidic Acid

  22. POST TRANSLATIONAL MODIFICATIONS • Loss Of N-formyl methionine or methionine • Acetylation of the amino group of amino terminal • Loss of SIGNAL SEQUENCES(15-30 aa at amino terminal) • ATTACHMENT OF CARBOHYDRATES ,PROTEOGLYCANS • ADDITION OF PROSTHETIC GROUPS eg HEME • PROTEOLYTIC PROCESSING egTrypsinogen to Trypsin • DISULFIDE CROSS LINKS eg Insulin

  23. The anticodon of yeast serinyl-tRNA is 5’-AGC-3’. Which of the following is most likely to be a serine codon? • 5’-ACG-3’ • 5’-CCG-3’ • 5’-GCU-3’ • 5’-UAG-3’ • 5’-UCG-3’

  24. Thank you

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