DNA replication.docx

DNA replicates via semi conservative replication- each strand of DNA serves as a template to synthesise a new complementary strand. Requirements for DNA replication Needs to be able to separate DNA and put them back together. Need to reproduce DNA sequence without error. DNA separation The hydr...

DNA replicates via semi conservative replication- each strand of DNA serves as a template to synthesise a new complementary strand. Requirements for DNA replication Needs to be able to separate DNA and put them back together. Need to reproduce DNA sequence without error. DNA separation The hydrogen bonds between the bases are broken via Heat and separates the double helix into 2 separate strands of DNA. Accuracy of DNA replication Is important hence steps such as: Proof reading- enzymes that reads the base sequence and removes any error in the sequence. Post-replication mismatch repair which reduced the errors further. Main enzymes involved in DNA replication are DNA polymerase I and III These enzymes are template dependent polymerases, can’t synthesise DNA strand by themselves whilst also requiring a primer. Primer Primers are complementary to DNA Short, single-stranded nucleic acid with free OH group at the 3’ end. Made up of RNA – RNA synthesis is required for DNA replication. Generated by enzymes called primase. Primases synthesises a short (5-10 bases) nucleotide sequence complementary to DNA. (primase doesn’t need a primer) After the primer is produced, DNA polymerase acts in elongation of the strand. Primer is removed after DNA synthesis by hydrolysis and replaced by DNA. The replication fork. Both strands of the DNA are synthesised at the same time and must be synthesised in the 5’ to 3’ direction. Replication fork is used to achieve the synthesises of new DNA. Structure of replication fork Consists of leading and lagging strand Consists of two branches, each one is made up of single strand DNA and each one serves as a template for DNA polymerase. Leading strand replication is continuous in 5’ to 3’ direction. Lagging strand replication is not continuous as the new DNA strand is synthesised in short fragments called Okazaki fragments- joined together by DNA ligase which catalyses the formation of phosphodiester bonds to form the new DNA strand. (ATP dependent process). Okazaki fragments are joined together within the duplex region. After the fragments are joined together, there are sometimes gaps which need to be filled by DNA polymerase I. RNA primer is also removed by DNA polymerase I as it contains exonuclease activity. Before replication, double stranded DNA needs to be unwound by enzyme called DNA helicase into single stranded DNA. Helicase uses ATP to separate DNA strands. The domains of helicase ‘pulls’ the DNA through the enzyme and the strands separate. As helicase separates the DNA helix, it causes the upstream DNA to become supercoiled due to stress on the structure. Supercoiling makes it harder for DNA to unwind but DNA needs to be relaxed to allow replication. This is achieved through action of topoisomerases enzyme. DNA replication is rapid and happens simultaneously on both the leading and lagging strands- this is achieved by dimerization of DNA polymerase. Origin of DNA replication In prokaryotes, the replication starts at origin of replication. Recognisable sequence within the genome- contains a lot AT tandem repeats which are essential for DNA replication and regulating replication. Also contains 5 copies of preferred binding site for dnaA protein. In eukaryotes, it is more complicated as genome is much bigger. Arranging DNA into linear chromosomes is also problematic for replication.