hapter 16 P. 305-324 16.1 Dna Is he enetic Material.H Morgan s group: showed that genes are located along chromosomes. wo chemical components of chromosomes are DN and protein. Little was known about nucleic acids. Role of DN in heredity was first worked out by studying bacteria and the viruses that infect them. Important Scientists in the Discovery of DN Frederick riffith Oswald very lfred Hershey and Martha hase Rosalind Franklin Francis rick and James Watson Frederick riffith Discovery of role in 1928 Vaccine against pneumonia (mice) Frederick riffith studied Streptococcus pneumoniae wo stains of the bacterium Pathogenic Non pathogenic Heated the pathogenic and killed the bacteria. Mixed the cell remains with living bacteria of the nonpathogenic and found some cells were then pathogenic Frederick riffith his newly acquired trait was inherited by all the descendants of the transformed bacteria. alled the phenomenon transformation: a change in genotype and phenotype due to the assimilation of external DN by a cell. 1
Oswald very Identity of transforming substance hree main candidates DN RN Protein very broke open the heat-killed bacteria and extracted the cellular contents Special treatments to inactivate each of the three molecules Oswald very ested each for its ability to transform live nonpathogenic bacteria. DN was left active transformation occurred ransforming agent was then announced as DN Studied viruses for more information Bacteriophages (phages): bacteria-eaters virus is composed of DN(or RN) enclosed by a protective coat. Fig. 16-4-3 Hershey and hase Devised an experiment showing that only one of the two components enters the E.coli cell. Specifically looked at 2 2 invades Escherichia coli bacteria Radioactive isotope of sulfur to tag protein, and phosphorus to tag DN. EXPERIMEN Phage Bacterial cell Batch 1: radioactive sulfur ( 35 S) Batch 2: radioactive phosphorus ( 32 P) Empty Radioactive protein protein shell DN Radioactive DN Phage DN entrifuge entrifuge Radioactivity (phage protein) in liquid Pellet (bacterial cells and contents) Radioactivity Pellet (phage DN) in pellet Rosalind Franklin Used X-Ray crystallography to find out structure of DN molecules X near center shows DN twists around ngle of the X suggests two strands and the nitrogenous bases are near the center of the molecule Shows diameter of the double helix Francis rick and James Watson Built three-dimensional models of DN Used Rosalind Franklin s x-ray pictures of DN to assist in the model he Double Helix Width suggested that it was made up of two strands. Began to build models that would conform to the X-ray measurements and the chemistry of DN. 2
Watson and rick- Double Helix omposed of two complementary strands of DN wrapped around each other Uniform diameter Hydrogen bonds held the two strands together wo hydrogen bonds between and hree hydrogen bonds between and. hargaff s Rule Studied percentages of nitrogenous bases. lmost equal % s denine bonds to hymine uanine bonds to ytosine = support Nitrogenous Bases make up DN molecules he two types are: Purines wo rings in the structure Pyrimidines One ring in the structure hargaff s Rule Fig. 16-5 Sugar phosphate backbone 5 end Nitrogenous bases hymine () denine () ytosine () Phosphate DN nucleotide Sugar (deoxyribose) 3 end uanine () Base Pairing Watson and rick stated their hypothesis Pair of templates, each of which is complementary to the other. Prior to duplication, the hydrogen bonds are broken he two chains unwind and separate Each chain acts as a template Eventually, two pairs of chains will result. 3
DN Replication In DN replication, the parent molecule unwinds, and two new daughter strands are built based on basepairing rules When a cell copies a DN molecule, each strand serves as a template for ordering nucleotides into a new, complementary strand. Nucleotides line up along the template strand and are linked Where there was one double-stranded DN molecule at the beginning, there are then two at the end. he copying mechanism is analogous to using a photographic negative to make a positive Watson and rick s Hypothesis Figure 16.9 (a) Parent molecule (b) Separation of strands (c) Daughter DN molecules, each consisting of one parental strand and one new strand Fig. 16-10 Parent cell First replication Second replication Replication Models Remained untested for many years Difficult to perform Watson and rick predicted the semiconservative model Each daughter molecule will have one old strand (derived or conserved from the parent molecule) and one newly made strand here are two others: onservative Dispersive (a) onservative model (b) Semiconservative model (c) Dispersive model DN Replication Models onservative he two parental strands reassociate after acting as templates for new strands. Semiconservative he two strands of the parental molecule separate, and each functions as a template for synthesis of a new, complementary strand. Dispersive Each strand of both daughter molecules contains a mixture of old and newly synthesized DN. Semiconservative Model 1950 Matthew Meselson and Franklin Stahl devised a clever experiment that supported the semiconservative model. Widely acknowledged among biologists to be a classic example of elegant experimental design. Figure 16.11 shows the experiment performed by Meselson and Stahl. 4
DN and Replication in Prokaryotes Prokaryotes: ring of chromosome holds nearly all of the cell s genetic material DN replication begins at a single point and continues to replicate whole circular strand Replication goes in both directions around the DN (begins with replication fork) Prokaryotes Eukaryotes Eukaryotic DN Replication he replication of a DN molecule begins at special sites called origins of replication Begins in hundreds of locations along the chromosome Begins when the DN molecule unzips creating: Replication fork Replication bubble Eukaryotic DN Replication Hydrogen bonds between base pairs breaks Helicases enzymes that untwist the double helix at the replication forks. Single-strand binding proteins bind to the unpaired DN strands, stabilizing them. opoisomerase relieves pressure of DN ahead of replication fork RN Primer RN chain Primase enzyme that synthesizes the primer Helicase will start to unwind the DN strand. opoisomerase will hold the strands together and prevent breaking Single-stranded binding proteins will stabilize the DN strands he Primase will start to form the RN chain Synthesizing a New DN Strand DN polymerase: catalyze the synthesis of new DN by adding nucleotides to a preexisting chain. Most DN polymerases require a primer and a DN template strand. DN polymerase III adds a DN nucleotide to the RN primer and then continues adding DN nucleotides complementary to the parent DN template strand. 5
ntiparallel Elongation he two strands of DN in a double helix are antiparallel (0riented in opposite directions). DN polymerases can only add nucleotides to the free 3 end of a primer or growing strand. NEVER HE 5 new strand can only elongate in the 5-3 direction LWYS Read in the 3 5 direction reated in 5 3 direction ntiparallel Elongation Leading Strand only 1 primer needed, moves toward the replication fork Lagging Strand many primers needed, moves away from the replication fork Okazaki Fragments on lagging strand, short segment of DN synthesized away from the replication fork DN ligase enzyme, joins the sugar-phosphate backbones of all the Okazaki fragments into a continuous DN strand he DN Replication omplex By interacting with other proteins at the fork, primase acts as a molecular brake, slowing progress of the replication fork. he DN replication complex does not move along the DN he DN moves through the complex Proofreading and Repairing DN During DN replication, DN polymerases proofread each nucleotide against its template as soon as it is added to the growing strand. he polymerase removes the incorrectly paired nucleotide and resumes synthesis. Mismatched nucleotides sometimes are missed. an also arise after replication Mismatched repair enzymes remove and replace incorrectly paired nucleotides that have resulted from replication errors. 6
Proofreading and Repairing DN Most cellular systems that repair incorrectly paired nucleotides use a mechanism that takes advantage of the base-paired structure of DN. Nuclease DN-cutting enzyme. uts out the segment of the strand containing the damaged segment. Enzymes involved in filling gaps: DN polymerase and DN ligase Nucleotide excision repair repair system, Figure 16.18 Replicating the Ends of DN elomeres Found at the ends of each chromosome and contain no genes (protective cap) elomerase lengthens telomeres in gametes dds DN bases at the 5 end he shortening of telomeres might protect cells from cancerous growth by limiting the number of cell divisions Important Enzymes to Remember Helicase, single-strand binding protein, topoisomerase Primase Synthesis of RN primer DN polymerase III (DN pol III) dd new bases to DN strand DN polymerase I (DN pol I) Removes and replaces RN primer from 5 end DN ligase Links Okazaki fragments and replaces RN primer from 3 end Vocab Nucleoid dense region of DN in a prokaryotic cell hromatin complex of DN and proteins that makes up a eukaryotic chromosome Heterochromatin Eukaryotic chromatin that remains highly compacted during interphase and is generally not transcribed. Euchromatin he less condensed form of eukaryotic chromatin that is available for transcription. hromosome Structures Bacteria: one double-stranded, circular DN molecule that is associated with a small amount of protein. Prokaryotes: Ring of chromosomes Holds nearly all the cell s genetic material Eukaryotes: DN in chromosomes Found in nucleus 7
Fig. 16-21a hromatin Packing In the cell, eukaryotic DN is combined with large amounts of protein. omplex of DN and protein chromatin Histones - proteins that are responsible for the first level of DN packing in chromatin Form a tight bond because DN is negatively charged and the histones have a positive charge DN double helix (2 nm in diameter) Histones Nucleosome (10 nm in diameter) Histone tail DN, the double helix Histones Nucleosomes, or beads on a string (10-nm fiber) H1 Fig. 16-21b hromatid (700 nm) hromosome Organizations 30-nm fiber Loops Scaffold 300-nm fiber 10-nm fiber 30 nm fiber 300-nm fiber Replicated chromosome (1,400 nm) 30-nm fiber Looped domains (300-nm fiber) Metaphase chromosome 10 - nm fiber DN winds around histones to form nucleosome beads Nucleosomes are strung together he string between the beads is called linker DN Nucleosome consists of DN wound twice around a protein core composed of two molecules each. 10-nm coils Forms a chromatin fiber 30 nm thick Interactions between nucleosomes cause the thin fiber to coil or fold into this thicker fiber 30-nm fiber 8
30 nm fiber forms loops called looped domains attached to a chromosome scaffold made of proteins Scaffold is rich in one type of topoisomerase. 300-nm Fiber Heterochromatin and Euchromatin Heterochromatin During interphase, a few regions of chromatin are highly condensed into heterochromatin Dense packing of the heterochromatin makes it difficult for the cell to express genetic information coded in these regions Euchromatin Most chromatin is loosely packed in the nucleus during interphase ondenses prior to mitosis DN ligase, DN polymerase, Helicase, Primase, elomerase Single-strand binding proteins, opoisomerase, Nuclease 1. removes section of DN that is damaged 2. proofreads and repairs damaged/mismatched DN; base pairing 3. synthesis of RN primer 4. Links Okazaki fragments; replaces RN primer from 3 end (in both leading and lagging strand). 5. relieves pressure of DN ahead of replication fork 6. attach to separated DN strands to ensure they stay separated 7. breaks hydrogen bonds between DN strands 8. lengthens telomeres in gametes Questions 9