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Chapter 3. the Beginnings of Genomic Biology – Molecular

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Chapter 3. the Beginnings of Genomic Biology – Molecular Chapter 3. The Beginnings of Genomic Biology – Molecular Genetics Contents 3. The beginnings of Genomic Biology – molecular genetics 3.1. DNA is the Genetic Material 3.6.5. Translation initiation, elongation, and termnation 3.2. Watson & Crick – The structure of DNA 3.6.6. Protein Sorting in Eukaryotes 3.3. Chromosome structure 3.7. Regulation of Eukaryotic Gene Expression 3.3.1. Prokaryotic chromosome structure 3.7.1. Transcriptional Control 3.3.2. Eukaryotic chromosome structure 3.7.2. Pre-mRNA Processing Control 3.3.3. Heterochromatin & Euchromatin 3.4. DNA Replication 3.7.3. mRNA Transport from the Nucleus 3.4.1. DNA replication is semiconservative 3.7.4. Translational Control 3.4.2. DNA polymerases 3.7.5. Protein Processing Control 3.4.3. Initiation of replication 3.7.6. Degradation of mRNA Control 3.4.4. DNA replication is semidiscontinuous 3.7.7. Protein Degradation Control 3.4.5. DNA replication in Eukaryotes. 3.8. Signaling and Signal Transduction 3.4.6. Replicating ends of chromosomes 3.8.1. Types of Cellular Signals 3.5. Transcription 3.8.2. Signal Recognition – Sensing the Environment 3.5.1. Cellular RNAs are transcribed from DNA 3.8.3. Signal transduction – Responding to the Environment 3.5.2. RNA polymerases catalyze transcription 3.5.3. Transcription in Prokaryotes 3.5.4. Transcription in Prokaryotes - Polycistronic mRNAs are produced from operons 3.5.5. Beyond Operons – Modification of expression in Prokaryotes 3.5.6. Transcriptions in Eukaryotes 3.5.7. Processing primary transcripts into mature mRNA 3.6. Translation 3.6.1. The Nature of Proteins 3.6.2. The Genetic Code 3.6.3. tRNA – The decoding molecule 3.6.4. Peptides are synthesized on Ribosomes CONCEPTS OF GENOMIC BIOLOGY Page 1 3.1. DNA IS THE GENETIC MATERIAL. (RETURN) CHAPTER 3. THE BEGINNINGS OF GENOMIC IOLOGY OLECULAR ENETICS (RETURN) B – M G In 1928, a British scientist, Frederick Griffith, published his work showing that live, rough, avirulent bacteria could be transformed by a “principle” found in dead, smooth, virulent bacteria into smooth, virulent As the development of classical genetics bacteria. This meant that the bacterial traits of rough proceeded from Mendel in 1866 through the early versus smooth and avirulence versus virulence were part of the 20th century the understanding that controlled by a substance that could carry the phenotype Mendel’s factors that produced traits were carried from dead to live cells. on chromosomes, and that there were infinite Griffith’s observations on Pneumococcus were ways that the genetic information from 2 parents controversial to say the least, and could assort in each generation to produce the inspired a spirited debate and genetic variety demanded by Darwin’s theories on much experimentation directed at “origin of species” on which natural selection proving whether the acted. This gave rise to the study of gene behavior “transforming principle” was of more complex traits and an understanding of protein or nucleic acid, the two genes in populations. main components of At the same time a quest for the material inside Frederick Griffith chromosomes identified early in (1879-1941) the 20th century, well before a cell, perhaps a subcomponent of a chromosome, Griffith’s experiments. This that carried the genetic instructions to make debate continued until Oswald Avery and his colleagues, organisms what they are was ongoing. Colin MacLeod, and Maclyn McCarty published their work in 1944 unequivocally showing that DNA was, in fact, Griffith’s transforming principle. This completely CONCEPTS OF GENOMIC BIOLOGY Page 2 revolutionized genetics and is considered the founding observation of molecular genetics. Figure 3.1. An electron micrograph of bacteriophage T2 (left), and a sketch showing the structures present in the virus (right). The head consists of a DNA molecule surrounded by proteins, while the core, sheath, and tail fibers are all made of protein. Oswald T. Avery Colin MacLeod Maclyn McCarty Only the DNA molecule enters the cell. In 1953, more evidence supporting DNA being the genetic material resulted from the work of Alfred Once it was established that DNA was the genetic Hershey and Martha Chase on E. coli infected with material carrying the instructions for life so to speak, bacteriophage T2. In their experiment, T2 proteins were attention turned to the question of “How could a labeled with the 35S radioisotope, and T2 DNA was molecule carry genetic information?” The key to that labeled with was labeled with the 32P radioisotope. Then became obvious with a detailed understanding of the the labeled viruses were mixed separately with the E. coli structure of the DNA molecule, which was developed by host, and after a short time, phage attachment was two scientists a Cambridge University, James Watson and disrupted with a kitchen blender, and the location of the Francis Crick. label determined. The 35S-labeled protein was found outside the infected cells, while the 32P-labeled DNA was inside the E. coli, indicating that DNA carried the information needed for viral infection. CONCEPTS OF GENOMIC BIOLOGY Page 3 Rosalind Franklin a young x-ray crystallographer working in the laboratory of Maurice Wilkins at 3.2. WASON & CRICK – THE STRUCTURE OF DNA. Cambridge University used a technique known as x-ray (RETURN) diffraction to generate images of DNA molecules that showed that DNA had a helical structure with repeating The basic laboratory observations that lead to the structural elements every 0.34 nm and every 3.4 nm formulation of a structure for DNA did not involve along the axis of the molecule. biologists. Rather Irwin Chargaff, an analytical, organic chemist, and physicists, Rosalind Franklin and Maurice Wilkins made the laboratory observations that led to the solution of the structure of DNA. Chargaff determined that there were 4 different nitrogen bases found in DNA molecules; the purines, adenine (A) and guanine G), and the pyrimidines, cytosine (C) and thymine (T), and he purified DNA from a number of different sources so he could examine the quantitative relationships of A, T, G, and C. He con- Rosalind Franklin Maurice Wilkins cluded that in all DNA molecules, the mole-percentage of A was nearly equal to the mole-percentage of T, while the mole-percentage of G was nearly equal to the mole- percentage of C. Alternatively, you could state this as the mole-percentage of pyri-midine bases equaled the mole- percentage of purine bases. These observations became Figure 3.2. X-ray diffraction image known as Chargaff’s rules. of DNA molecule showing helical structure with repeat structural elements. CONCEPTS OF GENOMIC BIOLOGY Page 4 These astute observations allowed Watson and Crick to synthesize together a 3-dimentional structure of a DNA molecule with all of these essential features. This structure was published in 1953, and immediately generated much excitement, culminating in a Nobel Prize in Physiology and Medicine, in 1962 awarded to Franklin, Wilkins, Watson, and Crick. Figure 3.4. Structures of purine and pyrimidine bases in DNA, and structure of 2’-deoxyribose sugar. The key elements of this structure are: Double helical structure – each helix is made from the alternating deoxyribose sugar and phosphate groups derived from deoxynuclotides, which are the monomeric units that are used to make up polymeric nucleic acid molecules. Each nucleotide in each chain Figure 3.3. Watson & Crick’s DNA structure. Their model consisted of consists of a nitrogen base of either the purine type a double helicical structure with the sugars and phosphates making (adenine or guanine) or the pyrimidine type (cytosine the two hlices on the outside of the structure. The sugars were held together by 3’-5’-phosphodiester bonds. The bases pair on the inside or thymidine) attached to the 1’-position of 2’- of the molecule with A always pairing with T, and G always pairing with deoxyribose sugar, and a phosphate group, esterified C. This pairing leads to Chargaff’s observations about bases in DNA. by a phospho-ester bond to the 5’-position of the sugar. CONCEPTS OF GENOMIC BIOLOGY Page 5 The nucleotides are held together in sequence order along the length of the polynucleotide chain by 3’-5’- Figure 3.5. The building bocks of nucleic phosphodiester bonds, and the strands demonstrate acids are nucleotides and nucleosides. Any a polarity as the 5’-OH at one end of a polynuc-leotide base together with a deoxyribose sugar forms a deoxyribonucleoside, while if the strand is distinct from the 3’-OH at the other end of sugar is ribose a ribonucleoside is formed the strand. Often, but not always, the 5’-strand end (not shown). Addition of a phosphate on the 5’ position of the sugar froms nucleotides will have a phosphate group attached. from nucleosides. Each of the 2 polynucleotide chains of the double helix are held together by hydrogen bonds beween the adenosines in one strand and the thymidines in the other strand, and between the guanosines in one strand hydrogen bonded to the cytosines in the other strand. Figure 3.7. Strand of DNA Figure 3.6. Base pairing between A and T involves two hydrogen showing the 3’,5’-phosphodi- bonds, and pairing between G and C involves 3 hydrogen bonds. ester bonds holding This means that the forces holding strands together in G=C base nucleotides together. pair-rich regions are stronger than in A=T base pair-rich regions. CONCEPTS OF GENOMIC BIOLOGY Page 6 In order to get a uniform diameter for the molecule and have proper alignment of the nucleotide pairs in the middle of the strands, the strands must be orient-ed in antiparallel fashion, i.e. with the strand polarity of each strand of the double helix going in the opposite direction (one strand is 3’-> 5’ whie the other is 5’ -> 3’).
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