Chapter 27 – Protein Metabolism27.1 The Genetic Code- Ribosomes are the site of protein synthesis from amino acids- Aminoacyl-tRNAs – tRNAs with an amino acid attachedo Aminoacyl-tRNA synthetases catalyze the addition of an amino acid totRNA- Translation – the overall process of mRNA-guided protein synthesisThe Genetic Code Was Cracked Using Artificial mRNA Templates- Codon – a triplet of nucleotides that codes for a specific amino acid- Reading frame – a new codon begins every three nucleotide residues- Initiation codon – AUG, the most common signal for the beginning of a polypeptide in all cellso AUG also codes for methionine- Termination codons – UAA, UAG, UGA, also called stop codons or nonsense codonso Signal the end of polypeptide synthesis and do not code for any aminoacids- Open reading frame (ORF) – a reading frame without a termination codon among 50 or more codonso Usually correspond to genes that encode proteins- Degenerate – an amino acid may be specified by more than one codonWobble Allows Some tRNAs to Recognize More than One Codon- When several different codons specify one amino acid, the difference between them usually lies at the third base position- Anticodon – a three-base sequence on the tRNA that base-pairs with mRNA codons- Wobble hypothesis :o The first two bases of an mRNA codon always form strong Watson-Crick base pairs with the tRNA anticodon and confer most of the coding specificityo The first base of the anticodon determines the number of codons recognized by the tRNA When the first base is C or A, base pairing is specific and only one codon is recognized When the first base is U or G, binding is less specific and the tRNA can recognize two different codons When inosine is the first nucleotide, three different nucleotidescan be recognizedo When an amino acid is specified by several different codons, the codons that differ in either of the first two bases require different tRNAso A minimum of 32 tRNAs are required to translate all 61 codons- Because the wobble base pairs loosely, it permits rapid dissociation of the tRNA from its codon during protein synthesisTranslational Frameshifting and RNA Editing Affect How the Code Is Read- Translational frameshifting – the reading frame is offset- RNA editing – the addition, deletion, or alteration of nucleotides in the RNA in a manner that affects the meaning of the transcript- ADARs – adenosine deaminases that act on RNA- APOBEC – apoB mRNA editing catalytic peptide family of enzymeso AID – activation-induced deaminase27.2 Protein SynthesisProtein Biosynthesis Takes Place in Five Stages- Stage 1: Activation of Amino Acidso Two requirements for synthesis of a polypeptide: The carboxyl group of each amino acid must be activated to facilitate formation of a peptide bond A link must be established between each new amino acid and the information in the mRNA that encodes it These are met by attaching the amino acid to a tRNA in the firststage of protein synthesis in the cytosolo Amino acids are attached to tRNA using ATP and Mg2+-dependent activating enzymes known as aminoacyl-tRNA synthetases- Stage 2: Initiationo The mRNA bearing the code for the polypeptide binds to the smaller ribosomal subunit and to the initiating aminoacyl-tRNA The large ribosomal subunit then binds to form the initiation complexo Initiation requires GTP and is promoted by cytosolic proteins called initiation factors- Stage 3: Elongationo The polypeptide is lengthened by covalent attachment of successive amino acid unitso Elongation requires elongation factorso The binding of each incoming aminoacyl-tRNA and the movement of the ribosome along the mRNA are facilitated by the hydrolysis of GTP- Stage 4: Termination and Ribosome Recyclingo Release factors aid the release of the polypeptideo Ribosome is recycled- Stage 5: Folding and Posttranslational Processingo The polypeptide may undergo enzymatic processing before or after folding Removal of one or more amino acids Addition of acetyl, phosphoryl, methyl, carboxyl, or other groups to amino acids Proteolytic cleavage Attachment of oligosaccharides or prosthetic groupsThe Ribosome Is a Complex Supramolecular Machine- The ribosome is a ribozyme- Ribosomes of eukaryotic cells are larger and more complex than bacterial ribosomesTransfer RNAs Have Characteristic Structural Features- tRNAs consist of a single strand of RNA folded into a precise three-dimensional structure- Most tRNAs have a guanylate residue at the 5’ end and all have CCA(3’) at the 3’ end. - Hydrogen-bonding pattern of tRNA forms a cloverleaf structure with four arms- Amino acid arm – the arm that can carry a specific amino acid esterified by its carboxyl group to the 2’ or 3’ hydroxyl group of the A residue at the 3’ end of the tRNA- Anticodon arm – the arm that contains the anticodon- D arm – contains the unusual nucleotide dihydrouridine- T C arm – contains ribothymidine and psuedouridineStage 1: Aminoacyl-tRNA Synthetases Attach the Correct Amino Acids to Their tRNAs- Each enzyme is specific for one amino acid- The reaction catalyzed by an aminoacyl-tRNA occurs in two steps:o Am enzyme bound intermediate, aminoacyl adenylate, is formedo The aminoacyl group is transferred from enzyme-bound aminoacyl-AMP to its corresponding tRNAo Two high-energy phosphate bonds are expended for each amino acid activated, rendering the reaction irreversible- Proofreading by Aminoacyl-tRNA Synthetaseso The aminoacylation of tRNA accomplishes two things: It activates an amino acid for peptide bond formation It ensures appropriate placement of the amino acid in a growing polypeptideo If available binding interactions do not provide sufficient discrimination between two substrates, the necessary specificity can be achieved by substrate-specific binding in two successive steps- Interaction between an Aminoacyl-tRNA Synthetase and a tRNA: A “Second Genetic Code”Stage 2: A Specific Amino Acid Initiates Protein Synthesis- All organisms have two tRNAs for methionineo One is used exclusively when (5’)AUG is the initiation codono The other is used to code for a Met residue in an internal position in a polypeptide- N-formylmethionyl-tRNAfMet (fMet-tRNAfMet) is formed in two reactions:o Methionine is attached to tRNAfMet by the Met-tRNA synthetaseo A transformylase transfers a formyl group from
View Full Document
Unlocking...