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Replication (14.3-14.6)

by: Feben Notetaker

Replication (14.3-14.6) BIOL 201L 004

Marketplace > Towson University > Biology > BIOL 201L 004 > Replication 14 3 14 6
Feben Notetaker
Introduction to Cell Biology and Genetics
Dr. Sarah Texel

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About this Document

Notes from the McGraw Hill text book and good for following lectures or extra study material.
Introduction to Cell Biology and Genetics
Dr. Sarah Texel
Class Notes
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This 8 page Class Notes was uploaded by Feben Notetaker on Monday October 19, 2015. The Class Notes belongs to BIOL 201L 004 at Towson University taught by Dr. Sarah Texel in Fall 2015. Since its upload, it has received 17 views. For similar materials see Introduction to Cell Biology and Genetics in Biology at Towson University.


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Date Created: 10/19/15
Section 143 Basic Characteristics of DNA Replication Tuesday October 13 2015 339 PM Requires a large number of cellular proteins Watson and Crick39s model of DNA suggested that the basis of copying genetic information is complementary One chain of DNA determines the sequence of its duplex ln replication the sequence of parental strands must duplicate daughter strands 0 One parental helix with two strands yields two daughter helices with four strands 0 Two daughter molecules are then separated during cell division Three models of DNA are possible 0 Conservative both strands of the parental duplex would remain intactconserved and new DNA copies would consist of allnew molecules Both daughter strands would be brand new 0 Semiconservative one strand of the parental duplex remains in daughter and an allnew complementary strand is built for the parental strand Daughter strands have one new and one parental strandquot 0 Dispersive copies of DNA would consist of mixtures of parental and newly synthesized strands new DNA is dispersed throughout each strand of both daughter and original strand after replication THE M ESELSONSTAHLEXPERIM ENT Grew bacteria in a heavy 15N isotope which became incorporated into the bacterial DNA After several generations bacterial DNA in 15N became denser than bacterial DNA grown in normal 14N Bacterial DNA in 15N wastransferred to MN and collected DNA at various time intervals 15N bacterial DNA were put in cesium chloride solutions It was spun at high speeds in ultracentrifuge Centrifuge pulls cesium ionsto the bottom because of its density and created a gradient for cesium concentration DNA strands floated or sank till it matched the density of cesium at the location of the gradient 0 0 rounds0min DNA was equal to density of 15N o 1 round20min DNA density decreased to be intermediate bw 15N and MN 0 2 rounds 40min two density classes of DNA were observed one intermediate and one 14N Replication Page 1 Conservative model isn39t consistent because DNA density change over timerounds Semiconservative model is consistent because 1 light strand an 1 heavy strand was found Rejected because two different densities of strands were found M echanism of DNA is semiconservative DNA is replicated by opening up a DNA helix and making copies of both strands to produce two daughter helices consisting of one old and one new strand COM PON ENTS OF DNA REPIJCATION Template building blocksto make the copy Nucleotides something to copy Polymerase Enzymesomething to do the copying Steps include o Initiation o Eongation 0 Termination Enzymes work together to assemble a new strand DNA polymerase o By matching existing DNA bases w complementary nucleotides 0 lJnks nucleotides together to make a new strand 0 synthesizes new 339 strands to existing strands o anthesize from 539 to 339 direction DNA polymerase requires a primer to begin synthesis they can39t begin wo a strand of RNA or DNA basepaired to the template RNA polymerase don39t require a primer 0 They synthesize primers Replication Page 2 Section 144 Prokaryotic Replication Wednesday October 14 2015 908 AM Replication begins at oriC and ends at the terminus OriC has repeated nucleotidesthat bind to an initiator protein 0 Also as and AT rich sequence that can open easily during initiation of replication I Has 2 H bondsto break in comparison to 3 H bonds in C G Replication goes bidirectionally form a unique origin to a unique terminus o Replicon DNA controlled by a replicon I A chromosome origin 1 replicon 0 Eco 0 DNA polymerase group of enzymesthat build new DNA strandsfrom template 0 They only synthesize from 539339 I DNA polymerase I on lagging strand replaces RNA primase with DNA I DNA polymerase II involved in repair process I DNA polymerase responsible for bulk synthesis of DNA 0 Nuclease breaks phosphodiester bonds bw nucleotides I Endonuclease DNA cut internally I Exonuclease DNA chew at the end El Goes from 339 539 to proofread DNA replication 0 There are 5 known polymerases but not all are active during replication UNWINDING OF DNA 0 Helicases Enzymesthat use energy from ATPto unwind DNA 0 When single strands are made their hydrophobic bases are exposed to water 0 Singlestrandbinding protein SSB coat hydrophobic bases 0 Supercoiling takes place during unwinding of DNA 0 EX rubber bands unwinding picture 0 Topoisomerase enzyme that preventsthat supercoiling 0 DNA gyrase specifictopoisomerase involved in DNA replication 0 Antiparallel characteristic of DNA causes DNA replication to occur in opposite directions 0 A primer needs to be added to initiate replication 0 Leading Strand synthesized in acontinuous form from a single primer I Replication occurstoward the replication fork o Lagging Strand synthesized in adiscontinuous form from more than one primer I Replication occurs away from replication fork I Okazaki fragments fragments formed in discontinuous strand SYNTHESIS o Replication fork partial opening of DNA helix to form two single strands with a forked appearance 0 Where most enzymes are located 0 DNA prim ase enzyme that synthesizes primers required by DNA polymerase during rep 0 An RNA polymerase that synthesizes short stretches of RNA that function as primers for polymerase Leading Strand 0 Single primer initiates replication 0 Extended continuously by DNA Pol Ill o If enzyme remains attatched to template it replicatesthroughout the whole circular DNA E coli chromosome 0 Processivity enzymes ability to remain attached to template 0 DNA Pol III has high processivity due to having a 3 subunit I 3 subunit provides a clamp structure to hold on to the template III AKA quotthe sliding clampquot I Also in Eukaryotes Daughter circular DNA Lagging Strand 0 Prim ase is needed to synthesize primers for each Okazaki fragment Acttvfm of Topoisomeaee ill 0 All RNA primers would be replaced with DNA flJ uswfgh o Fragments become stitched together 0 DNA Pol III synthesizes Okazaki fragments imw quot39etwrf o Removing of RNA and replacing DNA is done by DNA Pol l 7 Ew w 0 DNA pol l uses 539 to 339 exonuclease to remove primers then uses 539 to 339 polymerase to 5 WE g 4 25 in replace RNA primers Replication Page 3 httpiytimqoomviyM Kqu508i4 maxresdefaultipg Removing of RNA and replacing DNA is done by DNA Pol l O 0 DNA pol l uses 539 to 339 exonuclease to remove primers then uses 539 to 339 polymerase to replace RNA primers 0 DNA ligase sealsthe phosophodiester bonds byjoining Okazaki fragments to form complete strands TERM INATION 0 Occurs on the opposite side of the oriC on circular chromosome 0 Produces two daughter molecules intertwined together 0 They are soon unlinked by the DNA gyrase o Replisom e a macromolecule made up of all enzymes involved in DNA rep 0 Also thought of asa quotreplication organellequot o A loop isformed on the lagging strand so the lagging and leading strand can replicate simultaneously 0 The replisome is stationary while the DNA movesthroughout replication Replication Page 4 http lh4ggpht com Ve1WeOQE6Xc UXNch W 27LRM rOsQimgge thumb 25255B3825255Dpngim gm ax800 Polymerase Heticeee int Fill I Primase 39 I SEE ESDNA we strand w Syn thegs Laggingstrand synthesis 132 clam 39 V httpwwwumichedu ruotoloimagesresearchapplicationsimage1png Section 145 Eukaryotic Replication Wednesday October 14 2015 908 AM Two main factors 0 Larger amount of DNA organized into multiple chromosomes 0 Unear structure of chromosomes 0 New enzymes are needed for the ends of the chromosomes Eukaryotic replication has multiple origin s M ultiple origins shorten the time of DNA replication due to the length of chromosome in comparison to Ecoli M ultiple origins multiple replicons Origins aren39t sequencespecific like oriC 0 Recognition depends on chromatin structure as well as on sequence Number of origins change during development to increase or decrease rate of replication 0 Each origin is used once per cell cycle Enzymology of Eukaryotes is more complex Similar to bacteria but larger and more complex Initiation to assemble the helicases and primase complexes and loading the polymerase with its sliding clamp unit takes fore factors Eukaryotic primase is both an RNA polymerase and a DNA polymerase 0 First makes RNA primers then extends it with DNAto produce final primer Main replication polymerase hastwo enzymes that work together 0 DNA polymerase s 0 DNA polymerase 6 Proliferating cell nuclear antigen PCNA keeps sliding clamp complex together 0 First identified as an antibodyinducing protein in dividing cells 0 Similar to Bsubunit Same rep fork components Archaeal rep is more similar to eukaryotes than bacteria Main rep polymerase is most similar to eukaryotic pol 8 Sliding clamp is similar to the PCNA protein Specialized Ends Telomeres specialized structures found on the ends of eukaryotic chromosomes 0 Protect ends of chromosomesfrom nuclease and maintain integrity of Replication Page 5 linear chromosomes 0 Made of diff DNA sequences but not made by rep complex Replicating ends are complicated due to the linear structure in eukaryotes On the leading strand when DNA is being replicated the template runs out of nucleotides and the process is finished When the lagging strand reachesthe end of rep there is a gap left form the removal of the last primer It can39t be replaced and therefore can39t end 0 This would be a gradual of shortening chromosome after each cell division When sequence of telomeres was determined they were made of short repeated sequences of DNA Telomerase enzyme that has an internal RNA template that synthesizes the end of the DNAtemplate TELOM ERASE AGING AND CANCER Shortening ends of chromosomes occurs in absence of telomerase During embryonic development and childhood development in humans telomerase acrivity is high Low in adult somatic cells If cells divide as a function they have high telomerase activity Normal cells undergo only a specified number of divisions when grown in culture because of telomere length Cancer cells have activation of telomerase which helps maintain telomere length 0 Only on aspect of cancer cells Replication Page 6 Section 146 DNA Repair Wednesday October 14 2015 908 AM DNA polymerase have 339 to 539 exonuclease activity which proofreadsthe replicated DNA Even after the DNA is proofread error can still occur 0 Error correction mechanisms are there to look over proofread DNA Cells are constantly exposed to agents that can damage DNA Agents such as radiation from UV rays Xrays and chemicals in the environment Damaged DNA can be mutated mutagen Agentsthat increasesthe number of mutations above background levels DNA repair systems are made to restore damaged DNA 0 Reduces but doesn39t eliminate mutations TYPES OF REPAIR Soecific repair target a single kind of lesion in DNA and repair only that damage Nonspecific repair repair use a single mechanism to repair multiple kinds of lesions in DNA Specific Repair Photorepair repair for a damage in cell caused by UV light 0 Thymine dimer isformed in DNA 0 Two thymine bases covalently link together 0 Photolyase enzyme absorbs light in visible light and uses energy to cleave thymine dimer I Found in bacteria singlecelled eukaryotes to humans Nonspecific Repair Excision repair damaged region is removed and is replaced by DNA synthesis 0 ln Ecoli it39s accomplished by proteins Replication Page 7 encoded by the uvr A B Cgenes I Uvr genes have more sensitivity to UV rays Uses uvrABC complex 0 Three steps I 1 Recognition of damage V 2 Removal of damaged region 3 Resynthesized using info on undamaged strand asatemplate Two categories 0 Errorfree first resort and not as serious of a situation 0 Errorprone used asa last resort when cell is overwhelmed I Called quotSOSresponsequot in E coli Cells also repair damage that breaks DNA 0 Uses enzymes related to recombination during meiosis Replication Page 8


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