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Central Dogma.pdf

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University of Ottawa
Natalie Goto

PROTEIN SYNTHESIS Central Dogma · the central dogma states that genes are expressed in the phenotype or morphology of an individual via protein action · the central dogma is split into two parts – transcription and translation · the information to make a protein comes from a gene in the DNA molecule – referred to as “a template” or “master” · in order for one gene to direct the production of numerous essential proteins, copies of the “master” must be made · every copy will eventually amount to a specific polypeptide sequence · therefore, the central dogma is a sequence of events that begin with a genetic code and end with a protein · the template region of DNA is first “copied”, or transcribed into an mRNA, and then “converted” into a polypeptide chain, or translated Ribonucleic Acid (RNA) · like DNA, RNA is a carrier of genetic information · RNA differs from DNA in the following ways:  RNA contains a ribose sugar, not a deoxyribose sugar  instead of thymine, RNA contains uracil , which is still able to pair up with adenine  RNA is single-stranded, whereas DNA is double-stranded · when a gene is transcribed into messenger RNA, only a single-stranded complementary copy is made, where uracil replaces thymine · the three major classes of RNA molecules are messenger RNA (mRNA) – made by RNA polymerase II, transfer RNA (tRNA) – made by RNA polymerase III, and ribosomal RNA (rRNA) – made by RNA polymerase I · the mRNA length depends on the length of the gene that is transcribed – the longer the base pair sequence, the longer the mRNA · tRNAs (about 70-90 nucleotides long) transfer the appropriate amino acid to the ribosome to build a protein, as dictated by the mRNA transcript · the rRNA is a structural component of a ribosome that, together with proteins, forms the ribosome Transcription and Translation: An Overview · the three parts to transcription are initiation, elongation, and termination · transcription initiation begins with RNA polymerase binding to a promoter region on the DNA template strand, located near the beginning of the gene · transcription elongation is the process where RNA polymerase puts the appropriate ribonucleotides together, building the mRNA transcript as dictated by the genetic code on the template DNA · transcription termination takes place at a location shortly after the end of the coding region of the DNA – the RNA polymerase reaches a signal to stop transcribing · translation is also subdivided into initiation, elongation, and termination · translation initiation is when the ribosome recognizes a specific sequence on the mRNA and binds to that site · translation elongation is when the ribosome moves along the transcript mRNA six nucleotide bases at a time -- the first three bases are translated while the next three are prepared for translation, and each translated triplet codon codes for a specific amino acid that is brought to the ribosome by tRNA and strung together to make the polypeptide chain · translation termination is when the codon on the mRNA representing a “stop” signal is translated – the ribosome falls off and the polypeptide chain is released The Genetic Code · a sequence of three nucleotides are used to code for the 20 amino acids found in proteins · each triplet of nucleotides is called a codon 3 · the use of three nucleotides results in 4 = 64 different possible combinations · more than one codon can code for a single amino acid, which indicates a redundancy in the genetic code · the only start codon is AUG (methionine), and the three stop codons are UAA, UAG, and UGA Transcription Initiation · RNA polymerase binds to the segment of DNAthat is to be transcribed and opens the double helix · the binding site for the RNA polymerase is a region just before the coding region of the gene, called a promoter region · the promoter region is high in adenine and thymine bases – the recognition site for RNA polymerase · A and T bases possess 2 H-bonds between them, while C and G bases possess 3 H-bonds between them · therefore, less energy is required to unwind and break apart the H-bonds between A & T than would be required to break those between C & G Elongation · the mRNA is built in the 5’ to 3’ direction · a primer is not required by the RNA polymerase, as was the case for DNA polymerase in DNA replication · this means that elongation of the mRNA transcript immediately follows the initiation step · the promoter is not transcribed · the strand of the DNA that is transcribed is called the template strand · the complimentary strand of the DNA that is not transcribed is called the coding strand Termination · RNA polymerase recognizes the end of the gene when it comes across a terminator sequence · terminator sequences between prokaryotes and eukaryotes differ · at the point of termination, the mRNA transcript disassociates with the DNA template strand, and RNA polymerase is free to transcribe another gene Posttranscriptional Modifications · in eukaryotic cells, the primary transcript needs to undergo capping, tailing and base excision, before it can leave the nucleus · capping is the addition of a 5’ cap to the start of the transcript, consisting of 7-methyl guanosine, which in turn, forms a modified guanine nucleoside triphosphate · capping has two functions: o to protect the mRNA form digestive nucleases and phosphatases as it exits the nucleus and enters the cytoplasm o to initiate translation · in addition to the cap, a tail, of about 200 adenine ribonucleotides, known as a poly-A-tail, is added to the 3’ end of the transcript by an enzyme called poly-A-polymerase · the entire mRNA transcript consists of two regions – a coding region called exons, and a non-coding region called introns · the introns are interspread among the exons · the introns are removed so that their translation is prevented · particles made of RNA and proteins, called spliceosomes, remove the introns and join the exons together so that the transcript is one continuous coding gene · the introns stay inside the nucleus and are degraded into recycled nucleotides · the “primed” mRNA transcript then moves out of the nucleus and into the cytoplasm where it will be translated · there is no “quality control” mechanism for the mRNA strand · an incorrectly-made mRNA transcript will amount to a defective protein · however, so long as the original DNA template strand is correct, the multiple copies of mRNA transcripts will more than likely compensate for any mistake in one mRNA strand Translation The Ribosome · the assembly of amino acids to the chain occurs on the surface of ribosomes · ribosomes…. o hold the mRNA and fRNAs in proper position so that the codons are read accurately o catalyze the formation of the peptide bonds between adjacent amino acids o determine where translation starts and ends · all ribosomes are made of two subunits of unequal size · each subunit contains rRNA and ribosomal proteins · smaller subunits are flatter, larger subunits are hemispheric or rounded · ribosomes do not carry genetic information, nor do they work on specific mRNAs – they can bind and translate any mRNA made by the cell The Role of Transfer RNA (tRNA) · an intermediate “facilitator” that facilitates the match between the mRNA codon and its corresponding amino acid · there are 64 possible tRNAs – one for each codon · each tRNA has two important parts to it: o a triplet sequence of codons complementing the mRNA codon, called an anticodon o a binding site for an amino acid · as the tRNA anticodon pairs up with the mRNA codon, it carries its amino acid · for example, if the anticodon reads AUA, then the codon on the mRNA is UAU, the code for tyrosine · tRNAs are recycled once they release their amino acid to the previous amino acid · the 3’ end of the tRNA is the binding site for the amino acid · tRNA is a tertiary structure molecule that has an acceptor arm · all tRNA acceptor arms end with an ACCA – OH 3’ end · when tRNAs are attached to their amino acids they are charged – when they lack their amino acids they are uncharged · since all tRNA acceptor arms are all the same, special enzymes, called aminoacyl-tRNA synthetase enzymes, are required to link up specific amino acids to specific tRNA · there are 20 different aminoacyl-tRNA synthetase enzymes – each form an aminoacyl- tRNA complex with the tRNA and its amino acid · the energy that is accumulated by the complex in forming it, is transferred to the formation of the peptide bond between adjacent amino acids at the transcript Elongation of the Polypeptide Chain · the start AUG codon is recognized by the ribosome · the reason why it can differentiate between a start codon and an AUG codon in the middle of a coding sequence is that the 3’ end of the rRNA of the smaller subunit and the sequence of mRNA situated about 10 nucleotides in front of the initiator codon interact, which helps align the initiator codon of the mRNA with the anticodon of the tRNA initiator · AUG codes for methionine – the first amino acid in every protein · the tRNA has two binding sites – the A (aminoacyl) site and the P (peptidyl) site · the P site is entered only be an initiator tRNA as an aminoacyl-tRNA complex · the second aminoacyl-tRNA must then enter the A site as specified by the second codon · once both tRNAs are bound onto the ribosome, the enzyme peptidyl transferase (located in the large subunit) links the two amino acids together · the first tRNA releases its amino acid (met), which in turn, binds to the second amino acid (via peptidyl transferase) to become part of a peptidyl-tRNA-complex · then the initiator t
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