Translation

• What is RNA translation?

  Protein synthesis , amino acids are added to a polypeptidic chain according to the sequence of mRNA
• What are the characteristics of translation in prokaryotic cells?

  An expensive process (uses most of the energy) , requires more than 200 different macromolecules , translation is coupled to transcription
• What is a codon?

  Triplet of bases that codifies for an amino acid (64)
• What are characteristics of the genetic code?

  Triplets are organized in mRNA in a continuous way , do not overlap , first codon establishes the reading frame , is not ambiguous (each codon codifies for one and only one amino acid) , is degenerated (one amino acid is codified by several codons) , number of codons is related to the abundance of each amino acid in proteins , almost universal (UGA is a stop in humans)
• What are stop codons?

  Interrupt the message , in rando sequences are present in 1/20 ration , open reading frame (ORF): a coding sequence present in RNA with more than 500 codons with no stop codon
• WHat is the process of translation?

  In a processive , template directed , sequential way , 5-3 direction in mRNA ( amino to carboxi in protein) , 15 aa/s
• what are the RNAs involved in translation?

  rRNA: ribosome provides the catalytic environment

  tRNA: link or adaptor between amino acids and mRNA

  mRNA: carries the sequence
• What is the structure of rRNA?

  COmplex secondary structures and contribute to architecture of ribosomes
• What are the characteristics of ribosomes?

  place of translation , three trna binding sites: E(exit) , P (peptidyl) , A (aminoacyl)
• What is the structure of tRNA?

  A L shaped molecule made out of a single tRNA molecules , 75 nucleotides , 3 or 4 arms with modified bases
• How are codons and anticodons paired?

  Position in codon and anticodon are given in 5-3 direction

  Bases in codon positions 1 and 2 bind to anticodon according to complementarity rules , bases in codon position 3 and anticodon position 1 are wobble bases.

  
  

  Inosine (a nucleotide with a base of hypoxanthine) , maybe be present in anticodon 1 and can bind to U , A or C in codon 3 position
• How do bases in wobble positions bind?

  May bind with several bases , depending on which ones are involved ,makes wobble pairing weaker and helps to speed up translation, due to wobble pairing a single tRNA can recognize several codons
• What are the steps of translation?

  1) amino acid activation

  2) Initiation

  3) Elongation

  4) termination

  5) folding and post translational modifications
• How are amino acids activated?

  Activation consists of the binding of an amino acid to its tRNA , an atp consuming process , catalyzed by amino acyl tRNA synthases , low mistake rate and proof reading , bind to amino acid with amino acid arm according to the anticodon sequence

  
  

  Amino acid binds to enzyme which induces conformational chaneg , ATP can bind forming AMP , pyrophosphate is released and tRNA binds
• What is the process of initiation?

  First amino acid is always methionine , in prokaryotes is modified to N formylmethionine , special tRNA that is used only for the first methionine (in this case 2 trnas one to read AUG and other to inset methionine). 5' UTR region there are shine dalgarno sequences which interact with TRNA in subunits (places mrna in ribosome), initiation factor 1 blocks A site and IF 3 prevents binding of 50s subunit so mrna will interact with shine dalgano sequences ans AUG will be orientated in P position . TRNA is placed in P position by IF 2 GTP and GTP hydrolysis releases IFs and 50s subunit associates
• What is the process of elongation?

  EF-Tu-GTP binds and dellivers an aminoacyl trna to A site on the ribosome which forms a correct codon anticodon complex leading to activation of EF-Tu-GTPase activity , EF-Tus exchanges the nulceotides. Peptide bond formation (nucleophillic attack of amino N of aa in A on carbonyl C of aa in P) and nascent peptide gets bound to tRNA in A site. Ribosome moves three positions so A site gets to next codon , TRNA peptide is pushed from A to P site and unloaded trna shifts to E site , carried out by EF-G
• What occurs in termination?

  either RF1 or RF 2 recognize and bind to stop codons and RF 3 gtp facilitates binding of RF1 or RF 2 to ribosome , polypeptidic chain is transferred to water , hydrolysis of GTP results in dissociation of release factors
• What are the characteristics of prokaryotic translation?

  Fast process , transcription and translation occur simultaneously , polisomes
• What are the differences between prokaryotic and eukaryotic translation?

  in eukaryotes mRNA is separated from DNA and is transported out to cytosol while in prokaryotes it occurs all in the cytosol
• How does translation occur in eukaryotes?

  eIF1 blocks E site , eIF1A blocks A site , IF 3 prevents binding of 60s subunit , eiF2 gtp binds to trna a met and drives it to P site , binding of eIF5 GTP , eIF4 interact with 5' cap and drives the mRNA to 40s , codon anticodon pairing , GTP hydrolysis , dissociation of the eIF , association of the 60s subunit , initiation complex is completed
• how does protein folding occur?

  require chaperons: heat shock protein Hsp70 , chaperonines , protein disulfide ismoerases , prolil cis trans isomerases