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Translation is the cellular process that converts the language of nucleic acids contained within the mRNA molecule (A, U, G, C) into the language of amino acids in a synthesized protein (methionine, alanine, histidine, etc.). This translation process relies on a genetic code that converts nucleic acid sequence into amino acid sequence. Note that the genetic code that is translated into protein functions within the mRNA molecule, not in the DNA.
The mRNA molecule is read by the ribosome during translation as a group of three consecutive nucleotides, a so called triplet code. Each combination of three nucleotides within the mRNA sequence is a codon. In addition to the codons, the mRNA contains several RNA sequence features that play roles in translation (figure 11.1). These sequence features are listed from 5’ to 3’ along the mRNA molecule below:
There are 20 different types of amino acids typically incorporated in proteins; however, there are 64 possible combinations of four nucleotides (A, U, G, and C) in a triplet codon RNA sequence. As a result, there are more codons than amino acid possibilities (see figure 11.2). Thus, the genetic code is degenerate, meaning that more than one codon encodes a particular amino acid type. For example, the amino acid valine (val) is encoded by four codons (5’-GUU-3’, 5’-GUC-3’, 5’-GUA-3’, and 5’-GUG-3’) that differ only in the 3’-most base (wobble base). These four codons are called synonymous codons because they all encode valine.
The genetic code is also unambiguous, meaning that a particular codon sequence only encodes one type of amino acid. For example, the codon 5’-AAA-3’ always encodes the amino acid lysine (lys). Further, the genetic code is commaless, meaning that codons are read by the ribosome consecutively one triplet sequence after another, with no spacer nucleotides between codons. The codons are also nonoverlapping, meaning each nucleotide in the coding region of the mRNA is a member of only one codon. Finally, the genetic code is universal, meaning that the same genetic code is used in all organisms.
The polypeptide chains synthesized during translation have directionality (polarity); however, since the language of proteins is different than the language of nucleic acids, the labels 5' and 3' do not apply to the polarity of polypeptide chains. Instead, the amino acid encoded by the start codon (closer to the 5’ end of the mRNA) contains a free amino (NH3+) chemical group and thus is said to be the amino or N-terminus of the polypeptide chain. As the polypeptide chain is synthesized, peptide bonds are formed between the carboxyl group (COO-) of the growing amino acid chain and the amino groups of incoming amino acids (see figure 11.3). Peptide bond formation involves a condensation reaction that releases water as a product. The final amino acid added to a polypeptide contains a free carboxyl chemical group and is called the carboxyl terminal or C-terminal end of the polypeptide chain. The C-terminal end of the polypeptide corresponds to the codon immediately before the stop codon (closer to the 3’ end of the mRNA).
Transfer RNA molecules (tRNAs) act as adaptor molecules in translation. tRNAs function to:
There are many different types of tRNA molecules in a cell. Each tRNA type is encoded by a different gene in the genome and has a unique anticodon sequence. tRNA types are also distinguished by the amino acid carried. For example, tRNAval carries the amino acid valine, while tRNAphe carries the amino acid phenylalanine. tRNA molecules are example noncoding RNAs (ncRNAs), meaning that tRNA molecules themselves are untranslated; however, tRNAs function directly in the cell.
Even though different tRNA types (tRNAval RNAphe) have unique anticodons and carry unique amino acids, all tRNA molecules share similar structural features (figure 11.4):
The process of attaching an amino acid to the 3' end of a tRNA molecule is called charging. Charging tRNAs involves enzymes called aminoacyl-tRNA synthetases. Aminoacyl-tRNA synthetases have the following features:
The mechanism used by an aminoacyl-tRNA synthetase to charge a tRNA molecule has the following steps (see figure 11.5):
Most synonymous codons (codons that encode the same amino acid) have identical bases at the first two nucleotide positions (the 5’-most base and the middle base of the codon). These first two bases obey the AU/GC base pairing rules when forming hydrogen bonds with the tRNA anticodon. Degeneracy in the genetic code typically occurs at the 3'-most base within the codon; the 3’-most base in the codon does not have to form conventional base pairing interactions with the 5’-most base of the anticodon, leading to “wobble.” The rules that govern codon-anticodon at this wobble position interactions are called the wobble rules (see Table 11.1). For example, a U base in the wobble position of the codon (3'-most base) can form hydrogen bonds with A, G, or I in the wobble position of the anticodon (5'-most base). Similarly, a U in the wobble position of the anticodon (5'-most base) can form hydrogen bonds with either A or G in the wobble position of the codon (3'-most base).
Table 11.1 Wobble Rules Table.
A group of tRNA types, with slightly different anticodon sequences, which recognize the same mRNA codon sequence are called isoacceptor tRNAs. For example, suppose we have the codon 5’-UUU-3’ in the mRNA. tRNA molecules containing either 3’-AAA-5’, 3’-AAG-5’, or 3’-AAI-5’ anticodons can recognize this mRNA codon, according to the wobble rules (see Table 11.1). Note that these isoacceptor tRNAs are charged by the same aminoacyl-tRNA synthetase and therefore, bear the same amino acid (phenylalanine). Alternatively, a tRNA with the 3’-GGU-5’ anticodon can recognize two codons: 5’-CCA-3’and 5’-CCG-3’ according to the wobble rules. Both codons encode proline.
Why is this wobble phenomenon advantageous to cells? Wobble gives the cell tremendous flexibility in codon:anticodon interactions. A single mRNA codon can be recognized by more than one tRNA anticodon, and a single tRNA anticodon can bind to more than one mRNA codon. This flexibility allows translation to occur at a reasonable rate and allows a cell to save energy in the synthesis of tRNA molecules. Instead of assembling 61 different types of tRNA molecules (one type for each possible sense codon, minus the three stop codons), a cell can get away with synthesizing only 30-40 types of tRNA molecules, with some tRNAs recognizing multiple mRNA codon sequences.
The multisubunit ribosome is the central figure in the translation process. The ribosome binds to the mRNA, serves as a site for mRNA codon:tRNA anticodon recognition, breaks the covalent bond between the 3’ end of the tRNA and the amino acid, and synthesizes peptide bonds between amino acids.
A single bacterial cell is thought to contain approximately 10,000 ribosomes; these bacterial ribosomes are called 70S ribosomes. Note that the “S” designation of ribosomes correlates roughly with the overall three-dimensional shape of the ribosome component; the larger the molecule, the larger the “S” value. The 70S ribosome consists of the following subunits (see figure 11.6):
Eukaryotic cells have cytosolic ribosomes called 80S ribosomes. Mitochondrial and chloroplast ribosomes resemble prokaryotic 70S ribosomes. The 80S ribosomes in the cytosol synthesize most cellular proteins and consist of the following subunits (see figure 11.7):
Translation in prokaryotes and eukaryotes occurs in three stages (see figure 11.8):
The 5’-untranslated region (5’-UTR) of prokaryotic mRNAs plays a critical role in translation initiation, as the 5’-UTR contains the ribosome-binding site. Specifically, the 5’-UTR contains a recognition sequence (5’-AGGAGGU-3’) called the Shine-Dalgarno sequence that forms hydrogen bonds with a complementary sequence within the 16S rRNA of the small ribosomal subunit (30S) (see figure 11.9). Hydrogen bond formation between the Shine-Dalgarno sequence and the 16S rRNA promotes ribosome assembly. Additionally, the Shine-Dalgarno sequence positions the P site of the ribosome properly at the start codon to initiate polypeptide synthesis.
Translation initiation in the bacterium E. coli includes the following steps (see figure 11.10):
Translation initiation is more complex in eukaryotes than in prokaryotes. Key differences between eukaryotic and prokaryotic translation initiation include (see figure 11.11):
The process of translation initiation in eukaryotes is as follows:
Figure 9.11 - Translation initiation in eukaryotes.
Translation elongation in bacteria involves a series of cycles, one cycle per sense codon in the mRNA. At the beginning of each cycle, the tRNA molecule located within the P site of the ribosome is either attached to single amino acid (beginning of elongation cycle one) or a chain of amino acids (later elongation cycles; see figure 11.12). The A and E sites of the ribosome are empty. Each elongation cycle then proceeds as follows:
Translation elongation works similarly in eukaryotes; however, there are some minor differences:
Translation termination in bacteria occurs as follows (see figure 11.13):
Translation termination works essentially the same way in eukaryotes. One minor diference is the use of a single eukaryotic release factor (eRF) protein that recognizes all three stop codons. eRF cleaves GTP to terminate translation in eukaryotes.
In the electron microscope, multiple ribosomes (polyribosomes or polysomes) are often observed attached to a single mRNA transcript (see figure 11.14). Polysomes have been observed attached to both prokaryotic mRNA and eukaryotic mature mRNA transcripts.
Since prokaryotic cells lack nuclei, transcription and translation are coupled, meaning that translation can begin before transcription is completed. In eukaryotes, transcription and translation are uncoupled. Transcription and RNA modifications occur within the nucleus. Translation occurs later, after the mature mRNA has been transported from the nucleus to the cytoplasm.
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