BCOR 101 CHAPTER 7 TEXTBOOK NOTES
BCOR 101 CHAPTER 7 TEXTBOOK NOTES BCOR 101
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BCOR 101 CHAPTER #7 READING NOTES CHAPTER #7- DNA STRUCTURE AND REPLICATION SECTION 7.1- DNA IS THE HEREDITARY MOLECULE OF LIFE Hereditary- molecule is the molecular substance that carries and conveys the species’ genetic information Five essential characteristics of hereditary material o Localized to the nucleus and a component of chromosomes o Present in a stable form in cells o Sufficiently complex to contain the genetic information required to direct the structure, function, development, and reproduction of organisms o Able to accurately replicate itself so that daughter cells contain the same information as parental cells o Mutable, undergoing mutation at a low rate that introduces genetic variation and serves as a foundation for evolutionary change DNA first noted in 1869 by Friedrich Miescher whom isolated it from white blood cell nuclei in a mixture of nucleic acids and proteins he called nuclein 1870s microscopic studied identified fusiom of male and female nuclei during reproduction Chromosomes were observed in nuclei shortly after Nuceli of different species contain a different number of chromsomes Males and females contribute equal amounts of chromosomes in reproduction 1895 Edmund Wilson accurately recorded that eggs and sperm contribute the same number of chromosomes during reproduction o Proposed that inheritance may be effected by the physical transmission of a particular chemical compound from parent to offspring 1900s- Mendel’s hereditary principles were rediscovered and disseminated through the scientific community 1903- Wlater Sutton and Theodor Boveri accuratel described the paralles between homologous chromosomes and sister-chromatid separation during meiotic cell division and the inheritance of genes 1920- principle constituent of nuclein was identified as DNA and its basic chemistry was deciphered o Determined to be a polynucleotide consisting of four repaeating subunits (nucleotides) held together by covalent bonds o Nucleotides are adenine (A), thymine (T), cytosine (C) and guanine (G) 1923- DNA localized to chromosomes which made it a candidate for the hereditry material but it is not the sole constituent of chromosomes o Proteins are present in high concentrations as well as RNA in the nucleus and around the chromosomes as well as many other compounds such as carbohydrates and lipids These were all considered as an option for hereditary material at one point o It was noted the proetins had 20 different amino acids whereas DNA has only 4 nucleotides Suggested that the “20 letter alphabet” of protein could contain more information than the “4 letter alphabet” of DNA Against all these thoughts, 3 experiments were conducted between 1928 and 1952 combined to identify DNA as the hereditary material of organism Frederick Griffith studied strand of the bacteria pneumococcus which causes fatall pneumonia in mice Modern biology focuses on his first few pages of his report which provided indirect evidence that DNA is the molecule responsible for conveying hereditary characteristics in bacteria The S and R forms of pneumococcus occurred in for antigenic types of bacteria identified as I, II, III, and IV and each antigenic type elicits a different immune response from the mouse immune system as a result of the presence of several genetic differences Griffith’s most important observations are derived from the four injection tests that he performed using S and R bacterial strains of different antigenic types; following each injection test he was able to draw blood from the injected mice and culture the blood to identify the type of bacteria growing His first three injection results show that injecting the mice with S Strain bacteria produces illness and death, that injection of heat killed S strain bacteria does not induce illness and injection of an R strain does not produce illness Most significant result came when he injected a mixture of heat killed S3 strain and living R2 strained he found that the mice became ill and died from pneumonia; the blood cultures resulted from the dead mice revealed living S3 bacteria Griffin proposed that this outcome was not a result of a simple mutational event and that a molecular component that he called the transformation factor is responsible for transforming R2 into S3 Also proposed the transformation factor was a molecule that carries hereditary information Shortly after Griffith, Martin Dawson working with Oswald Avery developed an in vitro transformation procedure to mix living R cells with the purified extract of cellular material derived from heat killed S3 cells containing the transformation factor Biochemical assays were done which revealed that the S3 extract contained mostly of DNA along with small amounts of RNA and trace amounts of proteins, lipids, and polysaccharides Direct evidence that DNA was the transformation factor came from an experiment performed by Avery and his colleagues Colin MacLeod and Maclyn McCarty in 1944 Experiment identify the role of DNA in transformation by eliminating lipids, polysaccharides, protein, RNA and DNA one at a time from the S3 extract In introducing enzymes to get rid of each component, the final results revealed that transformation did not occur when DNase was added to DNA clearly indicating that transformation is blocked by the destruction of DNA Based on these observations Avery MacLeod and McCarty correctly concluded that DNA is the transformation factor and the probable hereditary material Evidence in a 1952 report by Alfred Hershey and Martha Chase show that DNA but not protein is responsible for bacteriophage infection of bacterial cells In their experiment, Hershey and Chase took advantage of an essential difference between the chemical composition of DNA and protein to confirm the hereditary of DNA Proteins contain large amounts of sulfur but almost no phosphorus and conversely DNA contains a large amount of phosphorus but no sulfur; Hershey and Chase grew phage cultures and different growth medium one containing sulfur to label protein another containing phosphorus to label DNA the researchers use radioactive labeled phages from each media to infect unlabeled host bacterial cells in parallel experiments After a short of a time each mixture was agitated in a blender to separate bacterial cells from the now empty phage cells, the relatively large bacteria cells were easily separated from the empty phage cells by centrifuge The heavier bacteria collected in a pellet at the bottom where the later empty phages remain suspended in the supernatant testing each fraction for radioactivity revealed that virtually all the phosphorus label associated with the newly infected bacterial cells and almost none with the empty phages on the other hand the sulfur labels were found in the phage fraction and only trace amounts are found associate in the bacterial pellet The results demonstrated that phage DNA but not phage protein is transferred to host bacterial cells and directs the synthesis of phage DNA and proteins, the assembly of progeny phage particles and ultimately the lysis of infected cells Experiment demonstrated the transformation factor identified previously by Griffith was DNA; it also showed that Avery, MacLeod and McCarty were correct in concluding that DNA is the hereditary material SECTION 7.2: THE DNA DOUBLE HELIX CONSISTS OF TWO COMPLEMENTARY AND ANTIPARALLEL STRANDS DNA nucleotide has three compnents o A deoxyribose sugar o One of four nitrogenous bases o Up to three phosphate groups Deoxyribose sugar o 5 carbons identified as 1‘,2’, 3’,4’, and 5’ o An oxygen atoms connects to the 1’ carbon and to the 4’ to form a pentose ring o 5’ carbon protrudes from the ring from the 4’ carbon o Nucleotide is attached to the 1’ carbon by a covalent bond o Hydroxyl group attached to the 3’ carbon o Phosphate molecule or chain is attached at the 5’ carbon o Hydrogen atms at 2’ carbon instead of a hydroxyl group Four nitrogeonous bases o Single ring pyrimidines Cytosine and thymine o Double ring purines Adenine and guanine Nucleotides part of a polynucleotide chain have one phosphate group at their 5’ carbon that forms the covalent phosphodiester bond with the adjacent nucleotide in the strand dAMP and dGMpP carry purines adenine and guanine while dCMP and dTMP carry the pyrimidines cytosine and thymine o these are deoxynucleotide monophosphates free DNA nucleotides not in a chain carry a string of 3 phosphates at the 5’ carbon and are identified as dATP, dGTP, dCTP, and dTTP o collectively known as deoxynucleotide triphosphates nucleotides are assembled into the nucleotide by enzyme DNA polymerase, which catalyzes the formation of the phosphodiester bond between the 3’ hydroxyl group of one nucleotide and the 5’ phosphate group of an adjacent nucleotide each polynucleotide chain has a sugar phosphate backbone consisting of alternating sugar and phosphate groups throughout its length DNA is most stable as a double helix and the two nucleotide strands have two rules they need to follow in their specific relationship o Arrangement of the nucelotides is such that the nucleotide bases of one strand are complementary to the corresponding nucleotide bases on the second strand o The two strands are antiparallel in orientation Complementary base pairing joins a pyrimidine from one strand to a purine on the other o Chemical basis of this is due to hydrogen bonding numbers between the bases of different strands Noncovalent bonds that form between the partial charges that are associated with hydrogen, oxygen and nitrogen atoms of nucleotide bases Antiparallel strand nature is essential to the formation of stable hydrogen bonds o Brings the partial charges of the complementary nucleotides in alignment to form hydrogen bonds In DNA, distance from axis of symmetry to the outer edge of the sugar phosphate backbone is 10 angstroms (1 nm) and the molecular diameter at any part of the helix is 20 angstroms ( 2nm) Tight packing of DNA bases in the duplex leads to base stacking which imparts a twist on the double helix Base pair stacking causes major and minor grooves which partially expose the nucleotides o Regions where DNA- binding proteins can most easily make direct contact with nucleotides along one or both strands of the double helix SECTION 7.3: DNA REPLICATION IS SEMICONSERVATIVE AND BIDIRECTIONAL Three attributes of DNA replication shared by all organisms o Each strand of the parental DNA molecule remains intact during replication o Each parental strand serves as a template directing the synthesis of complementary, antiparallel daughter strand o Completion of DNA replication results in the formation of two identical daughter duplexes, each composed of one parental strand and one daughter strand 3 proposed models of DNA replication o Semiconservative DNA replication- propsed that each daughter duplex contains one original parental strand of DNA and one complementary, newly synthesized daughter strands o Conservative DNA replication- predicts that one daughter duplex contains two strands of the parental molecule and the other contains two newly synthesized daughter strands o Dispersive DNA replication- predicts that each daughter duplex is a composite of interspersed parental duplex segments and daughter duplex segments Messelson-Stahl Experiment o Proved that the semiconservative model of replication was the correct model In bacteria, DNA replication is bidirectional from one origin site o Proved by the pulse chase labeling experiment performed by Joel Huberman and Arthur Riggs Autoradiograph evidence revealed multiple origin sites for DNA replication Eukaryotic replication origins initiate asynchronously during S pahse Eukaryotic DNA replication produces sister chromatids SECTION 7.4: DNA REPLICATION PRECISELY DUPLICATES THE GENETIC MATERIAL Bacterial, archaeal, and yeast DNA replication begins at specific locations the bind replication initiation proteins. Specific conserved sequences are found in bacteria, but replication initiation is directed by chromatin state in eukaryotes DNA replication begins with the synthesis of an RNA primer by primase, followed by the synthesis of leading and lagging DNA strands by DNA polymerase To complete replication. RNA primers are removed by DNA polymerase, and DNA segments are joined by DNA ligase DNA polymerases not only to replicate DNA but also proofread newly synthesized DNA for accuracy Eukaryotic and archaeal DNA replication proteins have a high degree of homology reflecting a shared common ancestry. Bacteria have analogous proteins, but are ancestrally more distant Eukaryotic chromosomes have repetitive sequences called telomeres at their ends that shorten with each replication in somatic cell cycles Telomerase is a ribonucleoprotein that synthesizes telomeric repeat sequences to maintain telomere length in germ-line and stem cells SECTION 7.5: MOLECULAR GENETIC ANALYTICAL METHODS MAKE USE OF DNA REPLICATION PROCESSES The polymerase chain reaction (PCR) is used to produce large numbers of copies to target DNA sequences Dideoxynucleotide DNA sequencing is used to determine the sequence of DNA fragments Next-generation and third-generation DNA sequencing are much faster and far cheaper methods that have paved the way for large numbers of genome sequencing projects and personal human genome sequencing VOCABULARY WORDS Bacteriophage- a virus whose host is a bacterium Base stacking- a phenomenon of DNA base-pair interaction that rotates the base pairs around a central axis of symmetry and imparts twisting to a double helix Bidriectional DNA replication- the standard method of DNA replication that synthesizes new DNA in both directions from a replication origin Clamp loader- a multiprotein complex that pairs with DNA polymerase and the sliding clamp during replication Consensus sequence- a nucleotide sequence in a DNA segement derived by comparing sequences of similar segments from other genes or organisms. The most commonly occurring nucleotides at each position compromise the sequence Deoxynucleotide 5’- monophosphate (dNMPs)- monophosphate forms of deoxynucleotides Deoxynucleotide 5’ – triphosphate (dNTPs)- thriphosphate forms of deoxynucleotides Dideoxy DNA sequencing- a method of DNA sequencing devised by Fred Sanger that uses a mixture of deoxynucleotide and Dideoxynucleotide triphosphates to selectively block DNA replication producing a ladder of partially synthesized DNA strands of different lengths Dideoxynucleotide triphosphates (ddNtPs)- rare DNA nucleotides absent oxygen molecules at the 2’ and 3’ carbons that are most commonly used in Dideoxynucleotide DNA sequencing DNA ligase- an enzyme active in DNA replication that joins together segments of a DNA stand by catalyzing formation of phosphodiester bonds DNA polymerase- the large multisubunit complex responsible for the synthesis of new strands of DNA during DNA replication or DNA repair DNA proofreading- the capacity of many types of DNA polymerase to utilize a 3’ to 5’ exonuclease activity to remove and replace mismatched or damaged nucleotides during replication DNA replication- the synthesis of new DNA strands by complementary base pairing of nucleotides in a daughter strand to those in a template strand Helicase- in DNA replication, the enzyme responsible for breaking down hydrogen bonds between complementary nucleotides of a DNA duplex. Unwinding of the strands occurs ahead of the advancing replication fork Lagging strand- in DNA replication, the discontinuously synthesized strand whose Okazaki fragments are ligated to complete new strand synthesis Leading strand- In DNA replication, the continuously synthesized strand Major groove- the larger of two grooves formed in the DNA sugar-phosphate backbone by the helical twist of the double helix and exposing certain base pairs Minor groove- the smaller of two grooves formed in the DNA sugar-phosphate backbone by the helical twist of the double helix and exposing certain base pairs Next-generation sequencing- high throughout massively parallel DNA sequencing by synthesis Okazaki fragments- a short segment of newly synthesized DNA that is part of a lagging strand that is ligated to other Okazaki fragments to complete lagging strand synthesis Origin of replication- the specific sequence at which DNA replication begins Polymerase chain reaction (PCR)- a laboratory method for controlled replication of a specific target sequence of DNA in successive cycle. Using teo short single-stranded primers that bind to sequences on opposite sides of the target sequence, exponential replication of the target sequence occurs Primase (DnaG)- the specialized RNA polymerase that synthesizes the RNA primer during DNA replication Proliferating cell nuclear antigen (PCNA)- in eukaryoutic DNA replication, the functional equivalent of the bacterial sliding clamp that adheres DNA polymerase to the template strand and drives its progression Replication bubble- a region of active bidirectional DNA replication containing replication forks on each end, an origin of replication in the middle, and the leading and lagging strands in each half of the bubble Replication fork- in DNA replication, the site of the replisome structure, and the site of synthesis of the leading strand and lagging strand DNA Replisome- the large molecular machine located at the replication fork that coordinates multiple reaction steps during DNA replication RNA primer- in DNA replication, the short single-stranded RNA segment synthesized by primase. The 3’ end of the RNA primer is used by DNA polymerase to begins synthesis of DNA Single-stranded binding protein- in DNA replication, a protein that adheres to each template strand following unwinding by helicase to prevent strand reannealing before the arrival of the replication fork Sliding clamp- in bacterial DNA replication, the multisubunit protein complex the joins with DNA polymerase to hold polymerase on the template and helps drive polymerase along the template Sugar-phosphate backbone- the alternating sugar (deoxyribose or ribose) and phosphate molecule pattern of nucleic acid strands formed by the formation of phosphodiester bonds linking nucleotides in the strand Supercoiled DNA- the superhelical twisting of covalently closed circular DNA Telomerase- the ribonucleoprotein complex whose RNA component provides a template used to synthesize repeating DNA segments that form chromosome telomeres Telomere- Repeating DNA sequences, synthesized by telomerase, at the ends of linear chromosomes of eukaryotes; contains dozens to hundreds of copies of specific short DNA sequence repeats that buffer the coding sequence of the chromosome from loss during successive cycles of DNA replication Topoisomerase- enzyme that relaxes DNA supercoiling by controlled strand nicking and rejoining
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