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18. Protein Synthesis in Eukaryotes.pdf

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Biology (Sci)
BIOL 200
Richard Roy

Naveen Sooknanan McGill Fall 2011 Protein Synthesis in Eukaryotes: Once in the cytoplasm, mRNA is able to react with a whole new set of cytoplasmic proteins which are factors in translation. Many of the proteins found in these mechanisms are GTP-binding and act as great switches which can be used to regulate translation levels of a protein The genetic code indicates which amino acid should be added onto a growing polypeptide chain by a series of three nucleotides on an mRNA molecule, known as the codon Along with these codons is a start codon, AUG, which codes for the initiation of translation and a stop codon, either UAA, UAG or UGA, which code for translational termination RNA plays three important roles in proteins synthesis: mRNAs play a role in the genetic code, i.e. dictating the amino acid sequence tRNAs act as adaptors which bring the required amino acids to the translation site with its anticodon rRNA forms a complex with many proteins to form the ribosome, which is a 2 subunit ribonucleic complex tRNA is an oddly shaped RNA molecule which undergoes extensive processing in order to be fully functional They are synthesized by Pol III within the nucleus They form a cloverleaf structure which contains a loop at the bottom with a 3 nucleotide long sequence known as the anticodon o This sequence is used to base pair with the codons of mRNA o In reality, they form an L shaped structure which is useful in its ability to attach to an amino acid An important processing of the tRNA occurs before it can be fully functional: o The 5 end sequence s removed o A short intron is removed o A CCA is added to the 3 end. This is used as an adaptor sequence for the binding of an amino acid, called the acceptor stem o Extensive base modification occurs all over the tRNA molecule These nucleotides are strange and are not found in normal RNA Aminoacyl tRNA synthetase is an enzyme which attaches the correct amino acid to the CCA 3 end of the tRNA o A high energy ester bond is created between the carbonyl group of the amino acid and the hydroxyl group of the tRNA o One of these reactions requires 1 ATP o Each aminoacyl tRNA synthetase is specific to 1 amino acid and to its corresponding tRNAs 1 Naveen Sooknanan McGill Fall 2011 o AA tRNA synthetase catalyzes this reaction by putting the ends of the tRNA and aa close to each other The modification of these tRNA molecules allow for non-conventional nucleotides to be present in the anticodon region. This allows for nonconventional base pairing to occur ONLY within the ribosome and ONLY during translation By simple math, there are 64 possible codons which can be found on mRNA; 61 if we remove stop codons Since there are only 20 amino acids, there must be multiple tRNAs which can carry the same amino acid o There are, therefore, sometimes several codon/anticodon pairings which will code for the insertion of the same amino acid into the polypeptide One of the irregular nucleotides, inosine, it responsible for some nonspecific base pairing which account for many wobble base pair possibilities There is, therefore, some leniency to the binding possibilities of tRNA to mRNA, but there is no leniency when it comes to the attachment of a particular aa to a tRNA rRNA is synthesized by Pol I inside the nucleolus, which is a small structure found within the nucleus rRNA accounts for almost 80% of all RNA in the cell, so it must be heavily transcribed to satisfy continuous demand o rDNA rRNA Transcription units of rRNA have been found on chromatin o rRNA genes are often found in repeated sequences in the genome and are synthesized in one large strand In every eukaryote, rRNA is transcribed into an 18S, a 5.8S and a 28S units o The introns and other noncoding regions may be slightly different, but the exons are highly conserved o The order of the exons in the gene is always 18 5.8 28 o The Svedberg (S) unit is a unit concerning both mass and size; it involves the location a molecule takes upon sedimentation centrifugation The rRNAs undergo significant modification and processing within the nucleolus o Since it is synthesised in one long strand, it must be cleaved into individual components o Extensive base modification on these units is also completed o This process is mediates by small nucleolar RNAs which are part of larger complexes called snoRNPs o 5S rRNA is NOT transcribed in the nucleolus; it arrives there by diffusion and does not undergo processing rRNA folds itself into highly conserved stem loop patterns rRNA is an integral part of translation because it complexes with protein to form the ribosome; the translation factory In prokaryotes: 2Naveen Sooknanan McGill Fall 2011 o 23S and 5S rRNA find to L1, L2 and L3 proteins, 31 in total, to form the 50S large ribosomal subunit o 16S rRNA complexes with 21 other proteins, S1, S2 and S3, to form the 30S small ribosomal subunit o The two subunits can form the 70S prokaryotic ribosome In eukaryotes: o 28S and 5.8S rRNA bind to 50 proteins, L1, L2 and L3 to form the 60S large ribosomal subunit o 18S rRNA and 33 proteins, S1, S2 and S3, bind to form the 40S small ribosomal subunit o Together, these form the 80S eukaryotic ribosome The ribosome structure is highly conserved among all organisms o Since prokaryotic ribosomes are so similar to those of eukaryotes, it seems that transcriptional machinery was perfected very early in evolutionary time Focusing on the eukaryotic 80S ribosome, it exists in equilibrium between an active (bound) and inactive (separated) state The transcriptionally active state is when the two subunits are bound together and locked in place until translation is completed When separated, these subunits need to be bound by eukaryotic initiation factors (eIFs) in order to be kept inactive o The
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