Chapter 28: DNA Replication, Repair & Recombination

DNA Replication, Repair & Recombination begins by examining the fundamental chemistry of DNA synthesis, where DNA polymerases catalyze phosphodiester bond formation in the 5' to 3' direction, strictly requiring a template and a primer to initiate the process. The text details the complex mechanics at the replication fork, distinguishing between the continuous synthesis of the leading strand and the discontinuous synthesis of the lagging strand via Okazaki fragments, which are subsequently joined by DNA ligase. A significant portion of the discussion focuses on the topological challenges of DNA unwinding, explaining how helicases separate strands using ATP hydrolysis and how topoisomerases manage supercoiling by altering the linking number (Lk), composed of twist and writhe. The distinction between Type I topoisomerases, which relax DNA by cutting one strand, and Type II topoisomerases (like bacterial DNA gyrase), which introduce negative supercoils by cutting both strands, is emphasized. The narrative then moves to the high coordination required for replication, describing the role of the DNA polymerase III holoenzyme, the sliding clamp that ensures processivity, and the trombone model of lagging strand synthesis. The text contrasts prokaryotic initiation at the single oriC locus involving DnaA with eukaryotic replication, which utilizes multiple origins, the Origin Recognition Complex (ORC), and strict cell cycle regulation. The unique problem of replicating linear chromosome ends in eukaryotes is solved by telomerase, a specialized reverse transcriptase carrying its own RNA template to synthesize telomeres. The chapter also addresses genomic stability, categorizing DNA damage from oxidizing agents, alkylating agents, and UV light (causing thymine dimers), and detailing specific repair pathways. These include proofreading exonuclease activity, mismatch repair, base-excision repair involving glycosylases and AP endonucleases, and nucleotide-excision repair utilized by the UvrABC complex. The connection between defective repair systems and diseases such as cancer—highlighting genes like p53 and conditions like Xeroderma pigmentosum—is explored alongside the Ames test for mutagenicity. Finally, the chapter covers genetic recombination, describing mechanisms like strand invasion facilitated by RecA and the formation and resolution of Holliday junctions, which are critical for repairing double-strand breaks and generating genetic diversity.