Lecture 5- DNA Transcription Spring 2023 PDF

DNA ORGANIZATION/DNA TRANSCRIPTION Dr. Joanne Sadier  Intended learning objectives ◦ Recall the structure of DNA ◦ Cite the different component of DNA and their characteristics ◦ Define what is central dogma ◦ Differentiate between euchromatin and heterochromatin ◦ List our curr...

DNA ORGANIZATION/DNA TRANSCRIPTION Dr. Joanne Sadier  Intended learning objectives ◦ Recall the structure of DNA ◦ Cite the different component of DNA and their characteristics ◦ Define what is central dogma ◦ Differentiate between euchromatin and heterochromatin ◦ List our current understanding of the genetic code ◦ Define the start and stop codons ◦ State the factors needed for transcription ◦ Differentiate between eukaryotic and prokaryotic transcription ◦ Identify the process of maturation of mRNA ◦ In this chapter, we focus on the various ways DNA is organized into chromatin, which in turn is organized into chromosomes. These structures have been studied using numerous approaches, including biochemical analysis as well as visualization by light microscopy and electron microscopy.  Central dogma ◦ Transcription ◦ Nucleotide sequence in one strand of DNA is used to construct complementary RNA sequence (mRNA). ◦ mRNA moves into cytoplasm → binds to ribosome ◦ Translation ◦ Synthesis of proteins is directed by information encoded in mRNA (genetic code). ◦ Consists of nucleotide triplets called codons Genetic code? Each triplet (codon) encodes for insertion of specific amino acid into growing protein chain © 2017 Pearson Education, Ltd. Our current understanding of the genetic code  How genetic code was discovered? the enzyme functions metabolically in bacterial cells to As a result, the order in which degrade RNA. However, in vitro, in the presence of high ribonucleotides are added is random, concentrations of ribonucleoside diphosphates, the reaction depending on the relative can be “forced” in the opposite direction, to synthesize RNA, concentration of the four as illustrated in the figure. ribonucleoside diphosphates present the mRNA they used in their cell-free protein-synthesizing in the reaction mixture. system was either UUUUUU... , AAAAAA... , CCCCCC... , or GGGGGG.... They tested each of these types of mRNA to see which, if any, amino acids were consequently incorporated into newly synthesized proteins. ◦ This technique took advantage of the observation that ribosomes, when presented in vitro with an RNA sequence as short as three ribonucleotides, will bind to it and form a complex similar to what is found in vivo. The triplet RNA sequence acts like a codon in mRNA, attracting a tRNA molecule containing a complementary sequence  Four codons specify valine (GUU, GUC, GUA, and GUG, in the bottom left corner), and they differ only by their third letter. In this case, all four letters in the third position specify valine. He postulated that hydrogen bonding at the third position of the codon–anticodon interaction would be less spatially constrained and need not adhere as strictly to the established base-pairing rules.  The Genetic Code Is Nearly Universal ◦ Between 1960 and 1978, it was generally assumed that the genetic code would be found to be universal, applying equally to viruses, bacteria, archaea, and eukaryotes. For example, cell-free systems derived from bacteria could translate eukaryotic mRNAs. ◦ However, several 1979 reports on the coding properties of DNA derived from mitochondria (mtDNA) of yeast and humans undermined the hypothesis of the universality of the genetic language. ◦ Most surprising was that the codon UGA, normally specifying termination, specifies the insertion of tryptophan during translation in yeast and human mitochondria.  What if? ◦ single mRNA may have multiple initiation points for translation. If so, these points could theoretically create several different reading frames within the same mRNA, thus specifying more than one polypeptide 5386 nucleotides, which should encode a maximum of 1795 amino acids= 6 protein? this small virus in fact synthesizes nine proteins consisting of more than 2300 amino acids Observations  The enzyme traverses the entire gene until eventually it encounters a specific nucleotide sequence that acts as a termination signal. The conservation of the promotor sequences during evolution attests to the critical nature of their role in biological processes. Such termination sequences, about 40 base pairs in length, are extremely important in prokaryotes because of the close proximity of the end of one gene to the upstream sequences of the adjacent gene In bacteria, both strong promoters and weak promoters have been discovered, causing a variation in time of initiation from once every 1 to 2 seconds to as little as once every 10 to 20 minutes. Mutations in promoter sequences may severely reduce the initiation of gene expression. Rho is a large hexameric protein that physically interacts with the growing RNA transcript, facilitating termination of transcription. In bacteria, groups of genes whose protein products are involved in the same metabolic pathway are often clustered together along the chromosome. In many such cases, the genes are contiguous, and all but the last gene lack the encoded signals for termination of transcription ◦ RNA polymerase II (RNAP II and Pol II) is a multiprotein complex that transcribes DNA into precursors of messenger RNA (mRNA) and most small nuclear RNA (snRNA) and microRNA. ◦ It is one of the three RNAP enzymes found in the nucleus of eukaryotic cells. (“int” for intervening ◦ Interestingly, it appears that somewhat different mechanisms exist for different types of RNA, as well as for RNAs produced in mitochondria and chloroplasts. ◦ However, in the studies of all other RNAs—tRNA in higher eukaryotes and rRNAs and pre-mRNAs in all eukaryotes— precise excision of introns is much more complex and a much more interesting story. ◦ Introns in eukaryotes are categorized into several groups: ◦ Group I: present in rRNA and it contains the enzymatic activity. They are called Ribozymes ◦ Group II: in mRNA and tRNA also involves 2 autocatalytic reactions. Let s watch how it works in real time ◦ https://www.youtube.com/watch?v=aVgwr0QpYNE