BMS 100 DNA Synthesis 1 (2023) PDF

Physiology Concept 1.17 Cell cycle – Post-learning 1: DNA synthesis 1 Dr. Hurnik BMS 100 Week 9 Video Link https://ccnm.ca.panopto.com/Panopto/Pages/Vie wer.aspx?id=068ce9f7-fc26-48d7-ac6bafbb000a170a Today’s Overview In-class Phases: G1, S, G2, M Checkpoints: Start transition, G2/M, metaphase-to-an...

Physiology Concept 1.17 Cell cycle – Post-learning 1: DNA synthesis 1 Dr. Hurnik BMS 100 Week 9 Video Link https://ccnm.ca.panopto.com/Panopto/Pages/Vie wer.aspx?id=068ce9f7-fc26-48d7-ac6bafbb000a170a Today’s Overview In-class Phases: G1, S, G2, M Checkpoints: Start transition, G2/M, metaphase-to-anaphase Checkpoint regulation Cell cycle regulation in the presence of growth factors Cell cycle regulation in the presence of unfavourable conditions CKIs, RB, p53 Cell survival Post-learning 1. DNA synthesis 1 2. DNA synthesis 2 3. Mitosis Cell cycle - intro The orderly sequence of events by which a cell duplicates its contents and divides in two § DNA Replication occurs is what phase of the cell cycle? A) G1 B) S C) G2 D) M Adapted from: https://upload.wikimedia.org/wikipedia/commons/3/38/0332_Cell_Cycle_With_Cyclins_and_Checkpoints.jpg DNA Replication The DNA double helix acts as a template for its own duplication. § Each daughter cell will inherit a DNA double helix containing 1 original strand & 1 new strand. This is called semi-conservative replication Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.2. Page 239 Steps of DNA synthesis DNA Replication occurs in 4 main steps: § 1. Strand separation § 2. Primer creation § 3. DNA replication § 4. Primer removal Step 1 – Strand separation If needed, review the structure of DNA § What bonds help contribute to its stability? In order for DNA synthesis to begin, the DNA double helix must the opened-up. § Two proteins are needed to open up the double helix 1. DNA Helicase Unwinds double helix What types of bond does it break? 2. Single stranded binding proteins Bind tightly and cooperatively to stabilize the single strand conformation Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.14. Page 247 Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.15. Page 247 Step 1 - Strand separation: replication fork Opening of the double helix creates the replication fork Adapted fromMolecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.7. Page 243 At the replication fork a multienzyme complex, DNA Polymerase will be used to synthesizes both new daughter strands of DNA. § However, DNA polymerase can only add nucleotides to an existing strand of DNA § Only works in 5’ to 3’ direction Step 2 – Primer creation DNA polymerase can only add nucleotides to an existing strand of DNA. § Ie - it can only elongate a strand of nucleic acid Therefore, a primer must be build first to serve as a base-paired chain on which to add new nucleotides. § This primer is made of RNA by an enzyme called DNA primase. Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.10. Page 245 Step 2 – Primer creation continued Since DNA can only be synthesized in the 5’ to 3’ direction, the replication fork has an asymmetric structure. § Leading strand: synthesized continuously § Lagging strand: synthesized discontinuously. The direction of nucleotide polymerization is opposite to the overall direction of DNA chain growth RNA primer Adapted from: Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.7. Page 243 Step 2 – Primer creation continued On the leading strand only 1 primer is required at the start of replication On the lagging strand a primer is needed for each Okazaki fragment § FYI – these primers are ~10 nucleotides long and are made at intervals of 100-200 nucleotides on the lagging strand Adapted from: Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.11. Page 245 Step 3 – DNA replication Once the primers are built, DNA polymerase can add nucleotides in the 5’ to 3’ direction Step 4 – Replacing the primers RNA primers are removed by a DNA repair system (FYI – RNAse H) and replaced with DNA. DNA ligase then joins the 3’ end of the new DNA fragment with the 5’ end of the previous fragment Adapted from: Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5.11. Page 245 What about supercoiling? As the replication fork moves along the double-strand DNA, anything in front of the replication fork will become overwound forming supercoils. Image adapted from: Kcneuman, CC BY-SA 4.0 , via Wikimedia Commons. Steps are FYI – to guide understanding only DNA topoisomerase The enzyme DNA topoisomerase relieves the super-helical tension by breaking the phosphodiester bond. This allows the two sections of the DNA helix to rotate freely & relieve tension. The phosphodiester bond will reform as DNA topoisomerase leaves. Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5-21. DNA synthesis Summary To do at home: § Label the diagram with the enzymes or proteins needed for each step. Molecular Biology of the Cell (Alberts et al) 6th ed. Figure 5-23. Page 254 Histones In addition DNA replication in Eukaryotes, histones must also be synthesize so that the newly replicated DNA can be packaged into nucleosomes § Histone synthesis also occurs during the S phase of the cell cycle Histone chaperones (aka chromatin assembly factors) assist formation of assembly of histone octomer & nucleosomes References Alberts et al. Molecular Biology of the Cell. Garland Science. Betts et al. Anatomy and Physiology (2ed). OpenStax Pathologic Basis of Disease(Robbins and Cotran) 10th ed.