Chapter 9 DNA Replication and Recombination errors 9 1 Genetic Information Must Be Accurately Copied Every Time a Cell Divides A Errors arise whenever information is copied the more times it is copied the greater number of 9 2 All DNA Replication Takes Place in a Semiconservative Manner A Semiconservative replication only one sequence of bases can be specified by each template Each of the original nucleotide strands remains intact despite no longer being combined with the original nucleotide strand a Three proposed models of replication conservative replication dispersive replication and semi conservative replication B Modes of Replication a Replicons individual units of replication each which contain a replication origin 1 Replication beings and continues until the entire replicon has been replicated 2 Theta replication takes place in bacteria and generates a structure that resembles a theta The double stranded DNA begins to unwind at the replication origin producing a sindle stranded nucleotide strand that serves as a template 3 The unwinding double helix generates a loop termed a replication bubble It may be at one or both ends of the bubble making it progressively larger 4 Replication fork the point of unwinding where the two single nucleotide strands separate from the double stranded DNA helix 5 Bidirectional replication if there are two replications forks one at each end of the replication bubble the forks proceed outward in both directions simultaneously unwinding and replicating the DNA until they eventually meet 6 If a single replication fork is present it proceeds around the entire circle to produce two complete circular DNA molecules each consisting of one old and one new nucleotide strand 7 Circular DNA that undergo replication have a single origin of replication C Requirements of Replication a A template consisting of single stranded DNA b Raw materials substrates to be assembled into a new nucleotide strand c Enzymes and other proteins that read the template and assemble the substrates into a DNA molecule 1 Because of the semiconservative nature of DNA replication a double stranded DNA molecule must unwind to expose the bases that act as a template for the assembly of new polynucleotide strands which are complementary and antiparallel to the template strands 2 New DNA molecules are synthesized from deoxyribonucleodise triphosphates dNTP each consisting of a deoxyribose sugar and a base attached to three phosphate groups 3 DNA are added to the 3 hydroxyl group of the growing nucleotide stand that e attacks the 5 phosphate group of the incomind dNTP 4 DNA synthesis is not spontaneously a DNA Polymerases the enzymes that synthesize DNA can add nucleotides ONLY to the 3 end of the growing strand so new DNA strands always elongate in the same 5 to 3 direction D Direction of Replication b Because DNA templates are antiparallel and strand elongation is always 5 3 if the synthesis on one template processed right to left then the other must be opposite c Continuous replication As the DNA unwinds the 3 5 direction template strand is exposed and allows the new strand to be synthesized continuously in the 5 3 direction Allowing a leading strand d The other template strand is exposed in the 5 3 direction and the synthesis proceeds in the 5 3 direction therefore opposite of the unwinding e Discontinuous replication short length DNA needs to be unwound before synthesis the replication machinery runs out of template It must start anew at the replication fork and proceed in the opposite direction until it runs into the preciously replicated segment making the lagging strand f Okazaki fragments the short lengths of DNA produced by discontinuous replication of the lagging strand They are linked together to create a continuous DNA molecule 9 3 The Replication of DNA Requires a Large Number of Enzymes and Proteins A Bacterial DNA Replication a Initiation b Unwinding c Primers 1 The circular chromosome of E coli has a single replication origin oriC 2 Indicator protein binds to oriC and causes a short section of DNA to unwind i This unwinding allows helicase and other single strand binding proteins to attach to the polynucleotide strand 1 DNA synthesis requires a single stranded template and double stranded DNA must be unwound before DNA synthesis can take place 2 DNA helicase breaks the hydrogen bonds that exist between the bases of the two nucleotide strands of a DNA molecule and begins to unwind the strand i ii iii The initiator protein first separates DNA strands at the origin providing a short stretch of single stranded DNA to which a helicase binds to the lagging strand template at each replication fork After DNA has been unwound the single stranded nucleotide chains have the tendency to form hyrdorgen bonds and stick back together Secondary structures also may form between complementary nucleotides on the same strand 3 Single strand binding proteins stabilize the single stranded DNA long enough for replication to take place They attach tightly to the exposed single stranded DNA and form tetramers that cover from 35 65 nucleotides 4 DNA gyrase a topoisomerase protein essential for the unwinding process It reduced they torsional strain that builds up ahead of the replication fork as a result of unwinding i It reduces torque by making a double stranded break in one segment of the DNA helix and then resealing the broken ends of the DNA 1 DNA polymerases cannot initiate DNA synthesis they require a primer an existing 3 OH group to get started 2 Primase synthesizes short stretched of nucleotides or primers to get DNA replication started i ii It synthesizes a short stretch of RNA nucleotides which provides a 3 OH group to which DNA polymerase can attach DNA nucleotides All DNA molecules initially have short RNA primers embedded within them these primers are later removed and replaced by DNA nucleotides d Elongation e DNA ligase iii iv On the leading strand a primer is only required at the 5 end of the newly synthesized strand On the lagging strand a new primer must be generated at the beginning of each Okazaki fragment 1 After DNA is unwound and a primer has been added DNA polymerases elongate the polynucleotide strand by catalyzing DNA polymerization 2 DNA polymerase III A large multiprotein complex that acts as the main workhorse of replication It synthesizes nucleotide strands by adding new nucleotides to the 3 end of a growing DNA molecule It s polymerase activity allows it to add
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