PDF: Resumão Aulas Biologia Molecular
Código genético O número de letras no código
Código de 1 base = 4 a possíveis Código de 2 bases = 42= 16 a possíveis Código de 3 bases = 43= 64 a possíveis
4 bases nitrogenadas diferentes
O código genético é redundante. O código é lido a partir de um ponto fixo.
As três fases de leitura possíveis na síntese de proteínas. Uma molécula de tRNA.
Figure 6-53. Wobble base-pairing between codons and anticodons. If the nucleotide listed in the first column is present at the third, or wobble, position of the codon, it can base-pair with any of the nucleotides listed in the second column. Thus, for example, when inosine (I) is present in the wobble position of the tRNA anticodon, the tRNA can recognize any one of three different codons in bacteria and either of two codons in eucaryotes. The inosine in tRNAs is formed from the deamination of guanine, a chemical modification which takes place after the tRNA has been synthesized. The nonstandard base pairs, including those made with inosine, are generally weaker than conventional base pairs. Note that codon-anticodon base pairing is more stringent at positions 1 and 2 of the codon: here only conventional base pairs are permitted. The differences in wobble base-pairing interactions between bacteria and eucaryotes presumably result from subtle structural differences between bacterial and eucaryotic ribosomes, the molecular machines that perform protein synthesis. (Adapted from C. Guthrie and J. Abelson, in The Molecular Biology of the Yeast Saccharomyces: Metabolism and Gene Expression, p. 487–528. Cold Spring Harbor, NewExpression, p. 487–528. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press, 1982.)
Estrutura de uma endonuclease de splicingde tRNA ancorada a um precursor de tRNA.
Figure 6-5. A few of the unusual nucleotides found in tRNA molecules. These nucleotides are produced by covalent modification of a normal nucleotide after it has been incorporated into an RNA chain. In most tRNA molecules about 10% of the nucleotides are modified
Figure 6-56. Amino acid activation. The two-step process in which an amino acid (with its side chain denoted by R) is activated for protein synthesis by an aminoacyl-tRNA synthetase enzyme is shown. As indicated, the energy of ATP hydrolysis is used to attach each amino acid to its tRNA molecule in a high-energy linkage. The amino acid is first activated through the linkage of its carboxyl group directly to an AMP moiety, forming an adenylated amino acid; the linkage of the AMP, normally an unfavorable reaction, is driven by the hydrolysis of the ATP molecule that donates the AMP. Without leaving the synthetase enzyme, the AMP-linked carboxyl group on the amino acid is then transferred to a hydroxyl group on the sugar at the 3′ end of the tRNA molecule. This transfer joins the amino acid by an activated ester linkage to the tRNA and forms the final aminoacyl-tRNA molecule. The synthetase enzyme is not shown in this diagram.
Figure 6-58. The genetic code is translated by means of two adaptors that act one after another. The first adaptor is the aminoacyl-tRNA synthetase, which couples a particular amino acid to its corresponding tRNA; the second adaptor is the tRNA molecule itself, whose anticodon forms base pairs with the appropriate codon on the mRNA. An error in either step would cause the wrong amino acid to be incorporated into a protein chain. In the sequence of events shown, the amino acid tryptophan (Trp) is selected by the codon UGG on the mRNA.
Figure 6-61. The incorporation of an amino acid into a protein. A polypeptide chain grows by the stepwise addition of amino acids to its C-terminal end. The formation of each peptide bond is energetically favorable because the growing C-terminus has been activated by the covalent attachment of a tRNA molecule. The peptidyl-tRNA linkage that activates the growing end is regenerated during each addition. The amino acid side chains have been abbreviated as R1, R2, R3, and R4; as a reference point, all of the atoms in the second amino acid in the polypeptide chain are shaded gray. The figure shows the addition of the fourth amino acid to the growing chain.
Figure 6-62. Ribosomes in the cytoplasm of a eucaryotic cell. This electron micrograph shows a thin section of a small region of cytoplasm. The ribosomes appear as black dots (red arrows). Some are free in the cytosol; others are attached to membranes of the endoplasmic reticulum. (Courtesy of Daniel S. Friend.)
Figure 6-63. A comparison of the structures of procaryotic and eucaryotic ribosomes. Ribosomal components are commonly designated by their “S values,” which refer to their rate of sedimentation in an ultracentrifuge. Despite the differences in the number and size of their rRNA and protein components, both procaryotic and eucaryotic ribosomes have nearly the same structure and they function similarly. Although the 18S and 28S rRNAs of the eucaryotic ribosome contain many extra nucleotides not present in their bacterial counterparts, these nucleotides are present as multiple insertions that form extra domains and leave the basic structure of each rRNA largely unchanged.
Figure 6-65. Translating an mRNA molecule. Each amino acid added to the growing end of a polypeptide chain is selected by complementary base-pairing between the anticodon on its attached tRNA molecule and the next codon on the mRNA chain. Because only one of the many types of tRNA molecules in a cell can base-pair with each codon, the codon determines the specific amino acid to be added to the growing polypeptide chain. The threestep cycle shown is repeated over and over during the synthesis of a protein. An aminoacyl-tRNA molecule binds to a vacant A-site on the ribosome in step 1, a new peptide bond is formed in step 2, and the mRNA moves a distance of three nucleotides through the small-subunit chain in step 3, ejecting the spent tRNA molecule and “resetting” the ribosome so that the next aminoacyl-tRNA molecule can bind. Although the figure shows a large movement of the small ribosome subunit relative to the large subunit, the conformational changes that actually take place in the ribosome during translation are more subtle. It is likely that they involve a series of small rearrangements within eachinvolve a series of small rearrangements within each subunit as well as several small shifts between the two subunits. As indicated, the mRNA is translated in the 5′-to-3′ direction, and the N-terminal end of a protein is made first, with each cycle adding one amino acid to the C-terminus of the polypeptide chain. The position at which the growing peptide chain is attached to a tRNA does not change during the elongation cycle: it is always linked to the tRNA present in the P site of the large subunit
Figure 6-6. Detailed view of the translation cycle. In the initial binding event (top panel) an aminoacyl-tRNA molecule that is tightly bound to EF-Tu pairs transiently with the codon at the A-site in the small subunit. During this step (second panel), the tRNA occupies a hybridbinding site on the ribosome. The codon-anticodon pairing triggers GTP hydrolysis by EF-Tu causing it to dissociate from the aminoacyl-tRNA, which now enters the A-site (fourth panel) and can participate in chain elongation. A delay between aminoacyl-tRNA binding and its availability for protein synthesis is thereby inserted into the protein synthesis mechanism. As described in the text, this delay increases the accuracy of translation. In subsequent steps, elongation factor EF-G in the GTP-bound form enters the ribosome and binds in or near the A-site on the large ribosomal subunit, accelerating the movement of the two bound tRNAs into the A/P and P/E hybrid states. Contact with the ribosome stimulates the GTPase activity of EF-G, causing a dramatic conformational change in EF-G as it switches from the GTP to the GDP-bound form. This change moves the tRNA bound to the A/P hybrid state to the P-site and advances the cycle of translation forward by one codon. During each cycle of translation elongation, the tRNAs molecules move through the ribosome in antRNAs molecules move through the ribosome in an elaborate series of gyrations during which they transiently occupy several “hybrid” binding states. In one, the tRNA is simultaneously bound to the A site of the small subunit and the P site of the large subunit; in another, the tRNA is bound to the P site of the small subunit and the E site of the large subunit. In a single cycle, a tRNA molecule is considered to occupy six different sites, the initial binding site (called the A/T hybrid state), the A/A site, the A/P hybrid state, the P/P site, the P/E hybrid state, and the E- site. Each tRNA is thought to ratchet through these positions, undergoing rotations along its long axis at each change in location. EF-Tu and EF-G are the designations used for the bacterial elongation factors; in eucaryotes, they are called EF-1 and EF-2, respectively. For each peptide bond formed, a molecule of EF-Tu and EF-G are each released in their inactive, GDP-bound forms. To be used again, these proteins must have their GDP exchanged for GTP. In the case of EF-Tu, this exchange is performed by a specific member of a large class of proteins known as GTP exchange factors.
Figure 6-71. The initiation phase of protein synthesis in eucaryotes. Only three of the many translation initiation factors required for this process are shown. Efficient translation initiation also requires the poly-A tail of the mRNA bound by poly-A-binding proteins which, in turn, interact with eIF4G. In this way, the translation apparatus ascertains that both ends of the mRNA are intact before initiating . Although only one GTP hydrolysis event is shown in the figure, a second is known to occur just before the large and small ribosomal subunits join.
Metionina N-formil-Metionina Em procariotos todos os polipeptídeos começam com N-formil-metionina.
Em eucariotos todos os polipeptídeos sintetizados no citosol começam com um resíduo de metionina.
Identificação do códon iniciador Procariotos –Sequência Shine Dalgarno
Eucariotos –a sequencia AUG mais próxima do terminal 5’ do mRNA geralmente serve como códon de início da síntese.
Tradução 5 Tradução 5
Tradução 6 Tradução 6
Figure 6-73. The final phase of protein synthesis. The binding of a release factor to an A-site bearing a stop codon terminates translation. The completed polypeptide is released and, after the action of a ribosome recycling factor (not shown), the ribosome dissociates into its two separate subunits.
Figure 6-75. A polyribosome. (A) Schematic drawing showing how a series of ribosomes can simultaneously translate the same eucaryotic mRNA molecule. (B) Electron micrograph of a polyribosome from a eucaryotic cell. (B, courtesy of John Heuser.)
Figure 6-76. The rescue of a bacterial ribosome stalled on an incomplete mRNA molecule.The tmRNA shown is a 363-nucleotide RNA with both tRNA and mRNA functions, hence its name. It carries an alanine and can enter the vacant A-site of a stalled ribosome to add this alanine to a polypeptide chain, mimicking a tRNA except that no codon is present to guide it. The ribosome then translates ten codons from the tmRNA, completing an 1-amino acid tag on the protein. This tag is recognized by proteases that then degrade the entire protein.
Figure 6-7. Incorporation of selenocysteine into a growing polypeptide chain.A specialized tRNA is charged with serine by the normal seryl-tRNA synthetase, and the serine is subsequently converted enzymatically to selenocysteine. A specific RNA structure in the mRNA (a stem and loop structure with a particular nucleotide sequence) signals that selenocysteine is to be inserted at the neighboring UGA codon. As indicated, this event requires the participation of a selenocysteine-specific translation factor.
Figure 6-78. The translational frameshifting that produces the reverse transcriptase and integrase of a retrovirus.The viral reverse transcriptase and integrase are produced by proteolytic processing of a large protein (the Gag-Pol fusion protein) consisting of both the Gag and Pol amino acid sequences. The viral capsid proteins are produced by proteolytic processing of the more abundant Gag protein. Both the Gag and the Gag-Pol fusion proteins start identically, but the Gag protein terminates at an in-frame stop codon (not shown); the indicated frameshift bypasses this stop codon, allowing the synthesis of the longer Gag-Pol fusion protein. The frameshift occurs because features in the local RNA structure (including the RNA loop shown) cause the tRNALeu attached to the C-terminus of the growing polypeptide chain occasionally to slip backward by one nucleotide on the ribosome, so that it pairs with a U codon instead of the UUA codon that had initially specified its incorporation; the next codon (AGG) in the new reading frame specifies an arginine rather than a glycine. This controlled slippage is due in part to a stem and loop structure that forms in the viral mRNA, as indicated in the figure. The sequence shown is from the human AIDS virus, HIV. (Adapted from T. Jacks et al., Nature331:280–283, 1988.)
As chaperonas moleculares direcionam o dobramento de muitas proteínas
Figure 6-79. Steps in the creation of a functional protein.As indicated, translation of an mRNA sequence into an amino acid sequence on the ribosome is not the end of the process of forming a protein. To be useful to the cell, the completed polypeptide chain must fold correctly into its threedimensional conformation, bind any cofactors required, and assemble with its partner protein chains (if any). These changes are driven by noncovalent bond formation. As indicated, many proteins also have covalent modifications made to selected amino acids. Although the most frequent of these are protein glycosylation and protein phosphorylation, more than 100 different types of covalent modifications are known.
Figure 6-81. The co-translational folding of a protein.A growing polypeptide chain is shown acquiring its secondary and tertiary structure as it emerges from a ribosome. The N-terminal domain folds first, while the C-terminal domain is still being synthesized. In this case, the protein has not yet achieved its final conformation by the time it is released from the ribosome. (Modified from A.N. Federov and T.O. Baldwin, J. Biol. Chem. 272:32715–32718, 1997.)
Figure 6-82. A current view of protein folding.Each domain of a newly synthesized protein rapidly attains a “molten globule” state. Subsequent folding occurs more slowly and by multiple pathways, often involving the help of a molecular chaperone. Some molecules may still fail to fold correctly; as explained shortly, these are recognized and degraded by specific proteases.
Figure 6-83. The hsp70 family of molecular chaperones.These proteins act early, recognizing a small stretch of hydrophobic amino acids on a protein's surface. Aided by a set of smaller hsp40 proteins, an hsp70 monomer binds to its target protein and then hydrolyzes a molecule of ATP to ADP, undergoing a conformational change that causes the hsp70 to clamp down very tightly on the target. After the hsp40 dissociates, the dissociation of the hsp70 protein is induced by the rapid re-binding of ATP after ADP release. Repeated cycles of hsp protein binding and release help the target protein to refold, as schematically illustrated in Figure 6-82.
Figure 6-84. The structure and function of the hsp60 family of molecular chaperones.(A) The catalysis of protein refolding. As indicated, a misfolded protein is initially captured by hydrophobic interactions along one rim of the barrel. The subsequent binding of ATP plus a protein cap increases the diameter of the barrel rim, which may transiently stretch (partly unfold) the client protein. This also confines the protein in an enclosed space, where it has a new opportunity to fold. After about 15 seconds, ATP hydrolysis ejects the protein, whether folded or not, and the cycle repeats. This type of molecular chaperone is also known as a chaperonin; it is designated as hsp60 in mitochondria, TCP-1 in the cytosol of vertebrate cells, and GroEL in bacteria. As indicated, only half of the symmetrical barrel operates on a client protein at any one time. (B) The structure of GroEL bound to its GroES cap, as determined by x-ray crystallography. On the leftis GroES cap, as determined by x-ray crystallography. On the leftis shown the outside of the barrel-like structure and on the righta cross section through its center. (B, adapted from B. Bukace and A.L. Horwich, Cell92:351–366, 1998.)
Figure 6-86. The proteasome.(A) A cut-away view of the structure of the central 20S cylinder, as determined by x-ray crystallography, with the active sites of the proteases indicated by red dots.(B) The structure of the entire proteasome, in which the central cylinder (yellow)is supplemented by a 19S cap (blue)at each end, whose structure has been determined by computer processing of electron microscope images. The complex cap structure selectively binds those proteins that have been marked for destruction; it then uses ATP hydrolysis to unfold their polypeptide chains and feed them into the inner chamber of the 20S cylinder for digestion to short peptides. (B, from W. Baumeister et al., Cell92:367–380, 1998. © Elsevier.)
Figure 6-87. Ubiquitin and the marking of proteins with multiubiquitin chains.(A) The threedimensional structure of ubiquitin; this relatively small protein contains 76 amino acids. (B) The C- terminus of ubiquitin is initially activated through its high-energy thioester linkage to a cysteine side chain on the E1 protein. This reaction requires ATP, and it proceeds via a covalent AMP-ubiquitin intermediate. The activated ubiquitin on E1, also known as the ubiquitin-activating enzyme, is then transferred to the cysteines on a set of E2 molecules. These E2s exist as complexes with an even larger family of E3 molecules. (C) The addition of a multiubiquitin chain to a target protein. In a mammalian cell there are roughly 300 distinct E2-E3 complexes, each of which recognizes a different degradation signal on a target protein by means of its E3 component. The E2s are called ubiquitin-conjugating enzymes. The E3s have been referred to traditionally as ubiquitin ligases, but it is more accurate to reserve this name for the functional E2-E3 complex.
Figure 6-8. Two general ways of inducing the degradation of a specific protein.(A) Activation of a specific E3 molecule creates a new ubiquitin ligase. (B) Creation of an exposed degradation signal in the protein to be degraded. This signal binds a ubiquitin ligase, causing the addition of a multiubiquitin chain to a nearby lysine on the target protein. All six pathways shown are known to be used by cells to induce the movement of selected proteins into the proteasome.