hapter 6 he Molecular Basis of Inheritance PowerPoint Lecture Presentations for Biology Eighth Edition Neil ampbell and Jane Reece Lectures by hris Romero, updated by Erin Barley with contributions from Joan Sharp opyright 008 Pearson Education, Inc., publishing as Pearson Benjamin ummings Overview: Life s Operating Instructions In 953, James Watson and Francis rick introduced an elegant double-helical for the structure of deoxyribonucleic acid, or DN DN, the substance of inheritance, is the most celebrated molecule of our time Hereditary information is encoded in DN and reproduced in all cells of the body his DN program directs the development of biochemical, anatomical, physiological, and (to some extent) behavioral traits Fig. 6- oncept 6.: DN is the genetic material Early in the 0th century, the identification of the molecules of inheritance loomed as a major challenge to biologists he Search for the enetic Material: Scientific Inquiry When. H. Morgan s group showed that genes are located on chromosomes, the two components of chromosomes DN and protein became candidates for the genetic material he key factor in determining the genetic material was choosing appropriate experimental organisms he role of DN in heredity was first discovered by studying bacteria and the viruses that infect them Evidence hat DN an ransform Bacteria he discovery of the genetic role of DN began with research by Frederick riffith in 98 riffith worked with two strains of a bacterium, one pathogenic and one harmless
When he mixed heat-killed remains of the pathogenic strain with living cells of the harmless strain, some living cells became pathogenic He called this phenomenon transformation, now defined as a change in genotype and phenotype due to assimilation of foreign DN Fig. 6- EXPERIMEN Living S cells (control) RESULS Living R cells (control) Heat-killed S cells (control) Mixture of heat-killed S cells and living R cells Mouse dies Mouse healthy Mouse healthy Mouse dies Living S cells Evidence hat Viral DN an Program ells In 944, Oswald very, Maclyn Mcarty, and olin MacLeod announced that the transforming substance was DN heir conclusion was based on experimental evidence that only DN worked in transforming harmless bacteria into pathogenic bacteria More evidence for DN as the genetic material came from studies of viruses that infect bacteria Such viruses, called bacteriophages (or phages), are widely used in molecular genetics research Many biologists remained skeptical, mainly because little was known about DN nimation: Phage Reproductive ycle Fig. 6-3 Bacterial cell Phage head ail sheath ail fiber DN 00 nm In 95, lfred Hershey and Martha hase performed experiments showing that DN is the genetic material of a phage known as o determine the source of genetic material in the phage, they designed an experiment showing that only one of the two components of (DN or protein) enters an E. coli cell during infection hey concluded that the injected DN of the phage provides the genetic information nimation: Hershery-hase Experiment
Fig. 6-4- Fig. 6-4- EXPERIMEN EXPERIMEN Phage Radioactive protein Phage Empty protein Radioactive shell protein Bacterial cell Bacterial cell Batch : radioactive sulfur ( 35 S) DN Batch : radioactive sulfur ( 35 S) DN Phage DN Radioactive DN Radioactive DN Batch : radioactive phosphorus ( 3 P) Batch : radioactive phosphorus ( 3 P) Fig. 6-4-3 EXPERIMEN dditional Evidence hat DN Is the enetic Material Phage Bacterial cell Batch : radioactive sulfur ( 35 S) Empty protein Radioactive shell protein DN Radioactive DN Phage DN entrifuge Radioactivity (phage protein) in liquid Pellet (bacterial cells and contents) It was known that DN is a polymer of nucleotides, each consisting of a nitrogenous base, a sugar, and a phosphate group In 950, Erwin hargaff reported that DN composition varies from one species to the next Batch : radioactive phosphorus ( 3 P) entrifuge Pellet Radioactivity (phage DN) in pellet his evidence of diversity made DN a more credible candidate for the genetic material nimation: DN and RN Structure Fig. 6-5 Sugar phosphate backbone end Nitrogenous bases hargaff s rules state that in any species there is an equal number of and bases, and an equal number of and bases hymine () denine () ytosine () Phosphate DN nucleotide Sugar (deoxyribose) end uanine () 3
Building a Structural Model of DN: Scientific Inquiry Fig. 6-6 fter most biologists became convinced that DN was the genetic material, the challenge was to determine how its structure accounts for its role Maurice Wilkins and Rosalind Franklin were using a technique called X-ray crystallography to study molecular structure Franklin produced a picture of the DN molecule using this technique (a) Rosalind Franklin (b) Franklin s X-ray diffraction photograph of DN Fig. 6-6a Fig. 6-6b (a) Rosalind Franklin (b) Franklin s X-ray diffraction photograph of DN Fig. 6-7 Franklin s X-ray crystallographic images of DN enabled Watson to deduce that DN was helical nm 3.4 nm end Hydrogen bond end he X-ray images also enabled Watson to deduce the width of the helix and the spacing of the nitrogenous bases he width suggested that the DN molecule was made up of two strands, forming a double helix end 0.34 nm (a) Key features of DN structure (b) Partial chemical structure end (c) Space-filling nimation: DN Double Helix 4
Fig. 6-7a end Hydrogen bond end Fig. 6-7b nm 3.4 nm 0.34 nm (a) Key features of DN structure end (b) Partial chemical structure end (c) Space-filling Watson and rick built s of a double helix to conform to the X-rays and chemistry of DN Franklin had concluded that there were two antiparallel sugar-phosphate backbones, with the nitrogenous bases paired in the molecule s interior t first, Watson and rick thought the bases paired like with like ( with, and so on), but such pairings did not result in a uniform width Instead, pairing a purine with a pyrimidine resulted in a uniform width consistent with the X-ray Fig. 6-UN Purine + purine: too wide Pyrimidine + pyrimidine: too narrow Purine + pyrimidine: width consistent with X-ray data Watson and rick reasoned that the pairing was more specific, dictated by the base structures hey determined that adenine () paired only with thymine (), and guanine () paired only with cytosine () he Watson-rick explains hargaff s rules: in any organism the amount of =, and the amount of = 5
Fig. 6-8 oncept 6.: Many proteins work together in DN replication and repair he relationship between structure and function is manifest in the double helix denine () hymine () Watson and rick noted that the specific base pairing suggested a possible copying mechanism for genetic material uanine () ytosine () he Basic Principle: Base Pairing to a emplate Strand Since the two strands of DN are complementary, each strand acts as a template for building a new strand in replication In DN replication, the parent molecule unwinds, and two new daughter strands are built based on base-pairing rules Fig. 6-9- (a) Parent molecule nimation: DN Replication Overview Fig. 6-9- Fig. 6-9-3 (a) Parent molecule (b) Separation of strands (a) Parent molecule (b) Separation of strands (c) Daughter DN molecules, each consisting of one parental strand and one new strand 6
Fig. 6-0 Parent cell First replication Second replication Watson and rick s semiconservative of replication predicts that when a double helix replicates, each daughter molecule will have one old strand (derived or conserved from the parent molecule) and one newly made strand ompeting s were the conservative (the two parent strands rejoin) and the dispersive (each strand is a mix of old and new) (a) onservative (b) Semiconservative (c) Dispersive Experiments by Matthew Meselson and Franklin Stahl supported the semiconservative hey labeled the nucleotides of the old strands with a heavy isotope of nitrogen, while any new nucleotides were labeled with a lighter isotope he first replication produced a band of hybrid DN, eliminating the conservative second replication produced both light and hybrid DN, eliminating the dispersive and supporting the semiconservative Fig. 6- EXPERIMEN Fig. 6-a Bacteria cultured in medium containing 5 N RESULS 3 ONLUSION onservative Semiconservative Dispersive DN sample centrifuged after 0 min (after first application) First replication 4 DN sample centrifuged after 40 min (after second replication) Second replication Bacteria transferred to medium containing 4 N Less dense More dense EXPERIMEN Bacteria cultured in medium containing 5 N RESULS 3 DN sample centrifuged after 0 min (after first application) 4 DN sample centrifuged after 0 min (after second replication) Bacteria transferred to medium containing 4 N Less dense More dense 7
Fig. 6-b ONLUSION onservative Semiconservative First replication Second replication DN Replication: loser Look he copying of DN is remarkable in its speed and accuracy More than a dozen enzymes and other proteins participate in DN replication Dispersive etting Started Fig. 6- Origin of replication Parental (template) strand Daughter (new) strand Replication begins at special sites called origins of replication, where the two DN strands are separated, opening up a replication bubble Doublestranded DN molecule wo daughter DN molecules Replication bubble Replication fork 0.5 µm eukaryotic chromosome may have hundreds or even thousands of origins of replication Replication proceeds in both directions from each origin, until the entire molecule is copied (a) Origins of replication in E. coli Bubble Double-stranded DN molecule Parental (template) strand Daughter (new) strand Replication fork 0.5 µm nimation: Origins of Replication wo daughter DN molecules (b) Origins of replication in eukaryotes Fig. 6-a Fig. 6-b Origin of replication Parental (template) strand Double-stranded DN molecule Daughter (new) strand Parental (template) strand Daughter (new) strand Doublestranded DN molecule wo daughter DN molecules Replication bubble Replication fork 0.5 µm Bubble Replication fork 0.5 µm wo daughter DN molecules (a) Origins of replication in E. coli (b) Origins of replication in eukaryotes 8
Fig. 6-3 t the end of each replication bubble is a replication fork, a Y-shaped region where new DN strands are elongating Single-strand binding proteins Primase Helicases are enzymes that untwist the double helix at the replication forks Single-strand binding protein binds to and stabilizes single-stranded DN until it can be used as a template opoisomerase corrects overwinding ahead of replication forks by breaking, swiveling, and rejoining DN strands opoisomerase Helicase RN primer DN polymerases cannot initiate synthesis of a polynucleotide; they can only add nucleotides to the end he initial nucleotide strand is a short RN primer n enzyme called primase can start an RN chain from scratch and adds RN nucleotides one at a time using the parental DN as a template he primer is short (5 0 nucleotides long), and the end serves as the starting point for the new DN strand Synthesizing a New DN Strand Enzymes called DN polymerases catalyze the elongation of new DN at a replication fork Most DN polymerases require a primer and a DN template strand he rate of elongation is about 500 nucleotides per second in bacteria and 50 per second in human cells Each nucleotide that is added to a growing DN strand is a nucleoside triphosphate dp supplies adenine to DN and is similar to the P of energy metabolism he difference is in their sugars: dp has deoxyribose while P has ribose s each monomer of dp joins the DN strand, it loses two phosphate groups as a molecule of pyrophosphate 9
Fig. 6-4 Sugar Phosphate New strand end end Base emplate strand end end end DN polymerase Pyrophosphate end ntiparallel Elongation he antiparallel structure of the double helix (two strands oriented in opposite directions) affects replication DN polymerases add nucleotides only to the free end of a growing strand; therefore, a new DN strand can elongate only in the to direction Nucleoside triphosphate end end Fig. 6-5 Overview long one template strand of DN, the DN polymerase synthesizes a leading strand continuously, moving toward the replication fork Parental DN Primer Overall directions of replication RN primer Sliding clamp DN poll III nimation: Leading Strand Fig. 6-5a Fig. 6-5b Overview Primer Parental DN RN primer Sliding clamp DN pol III Overall directions of replication 0
Fig. 6-6 Overview o elongate the other new strand, called the lagging strand, DN polymerase must work in the direction away from the replication fork he lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DN ligase emplate strand Overall directions of replication RN primer Okazaki fragment nimation: Lagging Strand Overall direction of replication Fig. 6-6a Fig. 6-6b emplate strand Overview Overall directions of replication Fig. 6-6b Fig. 6-6b3 emplate strand emplate strand RN primer RN primer Okazaki fragment
Fig. 6-6b4 Fig. 6-6b5 emplate strand emplate strand RN primer RN primer Okazaki fragment Okazaki fragment Fig. 6-6b6 able 6- emplate strand RN primer Okazaki fragment Overall direction of replication Fig. 6-7 he DN Replication omplex Single-strand binding protein Helicase Parental DN DN pol III Primer Primase Overview Overall directions of replication DN pol III DN pol I 4 DN ligase 3 he proteins that participate in DN replication form a large complex, a DN replication machine he DN replication machine is probably stationary during the replication process Recent studies support a in which DN polymerase molecules reel in parental DN and extrude newly made daughter DN molecules nimation: DN Replication Review
Proofreading and Repairing DN Fig. 6-8 DN polymerases proofread newly made DN, replacing any incorrect nucleotides In mismatch repair of DN, repair enzymes correct errors in base pairing DN can be damaged by chemicals, radioactive emissions, X-rays, UV light, and certain molecules (in cigarette smoke for example) In nucleotide excision repair, a nuclease cuts out and replaces damaged stretches of DN DN polymerase DN ligase Nuclease Replicating the Ends of DN Molecules Fig. 6-9 Ends of parental DN strands Limitations of DN polymerase create problems for the linear DN of eukaryotic chromosomes he usual replication machinery provides no way to complete the ends, so repeated rounds of replication produce shorter DN molecules Parental strand Last fragment Previous fragment RN primer Removal of primers and replacement with DN where a end is available Second round of replication New leading strand New lagging strand Further rounds of replication Shorter and shorter daughter molecules Fig. 6-0 Eukaryotic chromosomal DN molecules have at their ends nucleotide sequences called telomeres elomeres do not prevent the shortening of DN molecules, but they do postpone the erosion of genes near the ends of DN molecules It has been proposed that the shortening of telomeres is connected to aging µm 3
If chromosomes of germ cells became shorter in every cell cycle, essential genes would eventually be missing from the gametes they produce n enzyme called telomerase catalyzes the lengthening of telomeres in germ cells he shortening of telomeres might protect cells from cancerous growth by limiting the number of cell divisions here is evidence of telomerase activity in cancer cells, which may allow cancer cells to persist oncept 6.3 chromosome consists of a DN molecule packed together with proteins he bacterial chromosome is a doublestranded, circular DN molecule associated with a small amount of protein Eukaryotic chromosomes have linear DN molecules associated with a large amount of protein hromatin is a complex of DN and protein, and is found in the nucleus of eukaryotic cells Histones are proteins that are responsible for the first level of DN packing in chromatin In a bacterium, the DN is supercoiled and found in a region of the cell called the nucleoid nimation: DN Packing Fig. 6-a Fig. 6-b hromatid (700 nm) DN double helix ( nm in diameter) Nucleosome (0 nm in diameter) 30-nm fiber Loops Scaffold Histones Histone tail H 300-nm fiber DN, the double helix Histones Nucleosomes, or beads on a string (0-nm fiber) Replicated chromosome (,400 nm) 30-nm fiber Looped domains (300-nm fiber) Metaphase chromosome 4
hromatin is organized into fibers 0-nm fiber DN winds around histones to form nucleosome beads Nucleosomes are strung together like beads on a string by linker DN 30-nm fiber 300-nm fiber he 30-nm fiber forms looped domains that attach to proteins Metaphase chromosome he looped domains coil further he width of a chromatid is 700 nm Interactions between nucleosomes cause the thin fiber to coil or fold into this thicker fiber Most chromatin is loosely packed in the nucleus during interphase and condenses prior to mitosis Loosely packed chromatin is called euchromatin During interphase a few regions of chromatin (centromeres and telomeres) are highly condensed into heterochromatin Dense packing of the heterochromatin makes it difficult for the cell to express genetic information coded in these regions Histones can undergo chemical modifications that result in changes in chromatin organization For example, phosphorylation of a specific amino acid on a histone tail affects chromosomal behavior during meiosis Fig. 6- RESULS Fig. 6-UN ondensin and DN (yellow) Outline of nucleus ondensin (green) DN (red at periphery) Sugar-phosphate backbone Nitrogenous bases Hydrogen bond Normal cell nucleus Mutant cell nucleus 5
Fig. 6-UN3 Fig. 6-UN4 DN pol III synthesizes leading strand continuously Parental DN DN pol III starts DN synthesis at end of primer, continues in direction synthesized in short Okazaki fragments, later joined by DN ligase Primase synthesizes a short RN primer Fig. 6-UN5 You should now be able to:. Describe the contributions of the following people: riffith; very, Mcary, and MacLeod; Hershey and hase; hargaff; Watson and rick; Franklin; Meselson and Stahl. Describe the structure of DN 3. Describe the process of DN replication; include the following terms: antiparallel structure, DN polymerase, leading strand, lagging strand, Okazaki fragments, DN ligase, primer, primase, helicase, topoisomerase, single-strand binding proteins 4. Describe the function of telomeres 5. ompare a bacterial chromosome and a eukaryotic chromosome 6