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Chap 17 - Transcriptlation.docx

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University of Lethbridge
BIOL 1010
Brent Sellinger

th Reece eet al. 9 Ed. 1 Chapter 17 From Gene to Protein I. The Flow of Information in a Cell Transcription Translation DNA ▯ RNA ▯ Polypeptide This above relationship is often referred to as the Central Dogma DNA encodes the instructions to make an individual with a unique combination of traits. • review DNA & RNA structure (Chapter 5) How are these traits encoded in the DNA? • Garrod (1909) and Beadle and Tatum (1930s) suggested that certain mutants were the result of differences in enzyme activity • Beadle and Tatum studied nutritional mutants of the fungus Neurospora crassa • auxotrophic mutants - unable to grow on media lacking certain essential molecules normally synthesized by wild type strains • Formulated the "one gene - one enzyme" hypothesis that has been restated as the "one gene - one polypeptide" hypothesis How is a protein synthesized (i.e., gene expression)? The steps involved in protein synthesis (DNA ▯ RNA ▯ Polypeptide) include 1. Transcription • Synthesis of RNA (transcript) from DNA template • Transcript or messenger RNA (mRNA) carries the instructions for the assembly of the polypeptide. • Other types of RNA are also involved in the synthesis of protein (What are they?) 2. Translation • Synthesis of the polypeptide according to instructions encoded on the mRNA • There is a change in language (i.e., cell must translate the base sequence of the mRNA molecule into the amino acid sequence of a polypeptide) • The site of translation is the ribosome Ribosome - complex particles composed of proteins and rRNA that facilitate the linking of amino acids into polypeptides Reece eet al. 9 Ed. 2 Chapter 17 Differences in Protein Synthesis Prokaryotic cell - no compartmentalization - transcription and translation may be coupled or occur simultaneously. Eukaryotic cell - transcription occurs in the nucleus. mRNA translocated from nucleus to cytoplasm where translation occurs. mRNA is modified or processed before it leaves the nucleus. II. The Genetic Code • How does a 4 letter alphabet (such as found in DNA and RNA) encode for twenty different amino acids? With a 4 letter alphabet: Asingle letter word (4) can code for 4 amino acids Atwo letter word (4 x 4) can code for 16 amino acids Athree letter word (4 x 4 x 4) can code for 64 amino acids • Experiments verified that the flow of information from DNA to polypeptides is based upon a triplet code (i.e., three nucleotide word) • In the triplet code, three nucleotides (known as a codon) encode for a single amino acid. • Genetic information is encoded as a sequence of non-overlapping nucleotide triplets or codons e.g., a sequence of 900 nucleotides (nt) is required to code for 300 amino acids • Aseries of experiments conducted in the 1960's determined which amino acids the 64 triplets (codons) encoded • 61 of the codons code for amino acids (Figure 17.5) • AUG - codes for methionine as well as the start or initiation codon – it signals where translation starts • Three of the codons encode stop or termination codons (UAA, UAG, UGA) which signal where translation stops • open reading frame (ORF) - The genetic code is redundant (multiple codons may code for the same amino acid) but not ambiguous (the same codon does not code for more than one amino acid). Reece eet al. 9 Ed. 3 Chapter 17 The order and sequencing of the genetic code (reading frame) is important to its interpretation. For example: AUGAAAGCCGGGCCCGC  MetLysAlaGlyPro  AUGAAAGCCGGGCCCGC   ***LysProGlyPro   (where *** represents a stop signal) AUGAAAGCCGGGCCCGC    GluSerArgAlaArg  Notice that three different peptides are produced from the same RNA sequence when different reading frames are used. Each double stranded DNA has 6 reading frames – three on the top strand and another three on the bottom strand • The genetic code is nearly universal (there are a few exceptions) and this has enabled the development of many biotechnology applications (e.g., E. coli producing human insulin). • The universality of genetic code suggests that the genetic code arose early on in the evolution of life on Earth III. What is a Gene? • It is a discrete unit of heredity information consisting of a sequence of nucleotides encoding a specific polypeptide or an RNA molecule (i.e., rRNA, tRNA). • Genes contain the instructions but are not directly involved in the synthesis of polypeptides Schematic Diagram of a Gene th Reece eet al. 9 Ed. 4 Chapter 17 IV Transcription (Figure 17.7) • Only one strand of DNA is transcribed; this strand is called the template strand • The other strand is called the sense or nontemplate strand. Why is it called the sense strand? • RNA polymerases use the template strand to synthesize mRNA (5' ▯ 3') oProkaryotes have one type of RNA polymerase that synthesizes all types of RNA including mRNA oEukaryotes produce three types of RNA polymerases (I, II, and III) and RNA polymerase II is involved in mRNA synthesis Steps in Transcription 1. RNA polymerase binding and initiation of transcription • Specific DNA sequences mark initiation and termination sites for transcription • RNA polymerase binds to DNA regions called promoters (~100 to several hundred bp in length) • promoter regions contain the transcriptional recognition as well as initiation sites (i.e., TATA boxes in eukaryotes) for RNA polymerases. Promoters also determine which strand of DNA is the template strand. • In eukaryotic cells, RNA polymerase II also requires other proteins or transcription factors to aid it in recognizing and binding to the promoter region (Figure 17.8) • Once the RNA polymerase is bound to the promoter it begins to separate the DNA strands at the initiation site • ADNA sequence that is transcribed is called a transcriptional unit 2. Elongation of the RNA strand • The RNA polymerase untwists one turn of the double helix at a time, separates strands and adds nucleotides to the 3' end of the growing RNA molecule (~40 nt/s in eukaryotes) Compare RNA polymerase to DNA polymerase 3. Termination of transcription • In bacteria, transcription continues until the RNA polymerase reaches a termination sequence. Once the termination sequence is reached, the RNA polymerase detaches from the template and releases
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