Specific problems. The genetic code. The genetic code. Adaptor molecules match amino acids to mrna codons

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1 Tutorial II Gene expression: mrna translation and protein synthesis Piergiorgio Percipalle, PhD Program Control of gene transcription and RNA processing mrna translation and protein synthesis KAROLINSKA INSTITUTET Department of Cell and Molecular iology Medical Nobel Institute Highlights mrna translation What is mrna translation? Molecular mechanisms Components Medical relevance Regulation mrna functions as the intermediate between DNA and protein How does the information stored in a linear sequence of nucleotides in the mrna translate to a linear sequence of amino acids to obtain proteins? 1

2 Transcription Same language: 1dNTP 1rNTP Protein synthesis Different languages: 1rNTP 1aa Specific problems All cells must be able to translate the nucleotidebased language into proteins The translation can not be done letter by letter because there are 4 bases in the DNA/RNA and 20 different amino acids RNA translation The genetic code The genetic code mrna is read in codons (nucleotide triplets), t ex ATG Every codon specifies an amino acid or a STOP in the translation process The genetic code is universal The genetic code is redundant since many amino acids are specified by several codons. lack letters: codons (nucleotide triplets) Red letters: amino acids Adaptor molecules match amino acids to mrna codons How are nucleotides coupled with amino acids? The adaptor molecule is called transfer-rna (trna) 2

3 The 3D structure of trna is L-shaped and exhibits binding sites for both amino acids and codons The genetic code 3 end Amino acid binding site Antico don Codon binding site The Wobble Wobble effect in base pairing between codons and anticodons lack letters: codons (nucleotide triplets) Red letters: amino acids A specific enzyme loads trna molecules with an amino acid The enzyme is called amino acyl t-rna synthetase 1) 2) The genetic code is translated with the help of two different adaptors The new amino acids are incorporated in the growing polypeptide chain Amino acyl trna Net result: one amino acid is chosen by one codon 3

4 A Ribosomes Where does translation take place? C Eukaryotic ribosome Prokaryotic ribosome Ribosomes Ribosomes are very complex catalytic machineries containing more than 50 different proteins as well as ribosomal RNA (rrna) Active ribosomes conist of a large and a small subunit Ribosomes are very conserved between prokaryotic and eukaryotic organisms In eukaryotes, active ribosomes are only present in the cytoplasm Antibiotics and mrna translation Antibiotics are designed against processes and structure which are unique for microbes and are not found in eukaryotes There are slight differences between eukaryotic and prokaryotic ribosomes and these differences are targeted by antibiotics There are antibiotics (eg. kanamycin) which specifically block bacterial ribosomes Protein synthesis is a dynamic process How can the ribosome facilitate and coordinate all the molecular mechanisms that take place for efficient mrna translation? 4

5 Ribosomes contain four binding sites trna binding sites There are three main steps in mrna translation inding of amino acyl-trna to A-site The C-terminus end of the peptide in the P-site is coupled to the N-terminus of amino acid in the A-site mrna binding site Conformational changes cause the shift of the mrna by exactly three nucleotides so that a new A-siteA becomes available Translation of an mrna molecule...require the help of elongation factors A D Peptidyl transferase Conformational changes in the mrna C E F Initiation and termination of translation requires specific signals Initiation requires a specific AUG codon, which exclusively binds to a trna carrying a methionine Translation is terminated when stop codons (UAA, UAG, UGA) are encountered by the ribosome None of the trna molecules recognizes stop codons Initiation requires initiation factors Initiator trna binds the small ribosomal subunit The small ribosomal subunit recognizes the 5 -cap structure in the mrna (it also needs initiation factors eif4e and eif4g) The small ribosomal subunit scans the mrna until it reaches the AUG codon The large ribosomal subunit binds the small ribosomal subunit to reconstitute an active ribosome. The initiation factors are dissociated 5

6 Initiation of translation Termination of translation A D E The proteins which are ready leave ribosomes C F The ribosomal subunits dissociate Proteins are made in polyribosomes The synthesis of most proteins takes between 20 seconds and few minutes A Quality control of newly synthesized proteins? Proteins are folded co-translationally Proteins acquire their secondary structure during the translation process Several mechanisms garantee correct protein folding 6

7 Abnormally folded proteins can aggregate and give rise to disease A long way from DNA to protein Prion, Huntington and Alzheimer are the result of accumulation of wrongly folded proteins which build filaments that can not be degraded and impair cellular function 7

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