section Ea.ppt

Electron Microscopy of replicating DNA reveals replicating bubbles Priming is required to start DNA synthesis The cell cycle is controlled by many cyclin-cdk complexes G1 S G2/M Cyclin-Cdk Cyclin-Cdk Cyclin-Cdk Budding Yeast CLN1,2,3-CDC28 CLB5,,(3,4)-CDC28 CLB1,2(3,4)-CDC28 Fission Yeast CIG1-CDC2 CIG2-CDC2 CIG13-CDC2 Higher Eukaryotes CyclinD1,2,3-CDK4/6 CyclinA-CDK2 CyclinB-CDC2 CyclinE1,2-CDK2 The Rb-E2F pathway Inhibitory control of cell cycle E4: Eukaryotic DNA replication Small animal viruses (simian virus 40猿猴病毒40, 5 kb) are good mammalian models for elongation (replication fork). 2. Yeast (Saccharomyces酵母菌cerevisiae啤酒): 1.4 x 107 bp in 16 chromosomes, 400 replicons复制子, good for studying replicons复制子 Experimental systems 3. Cell-free extract prepared from Xenopus非洲爪蟾 (frog) eggs containing high concentration of replication proteins and can support in vitro replication. Elongation Removal of RNA primer (引物), and gap filling with DNA pol I去除引物，再由DNA pol I将间隙填充 Ligation of Okazaki fragments (冈崎片段) are linked by DNA ligase (连接酶). 连接冈崎片段是通过DNA连接酶进行的 Elongation: lagging strand (滞后链) replication Polymerase III holoenzyme全酶 (DNA pol III) DNA pol I (5’?3’ exonulclease activity)去除引物 DNA pol I (5’?3’ polymerase activity) DNA ligase (NAD+为能量来源) 5’?3’ 5’?3’ 3’ → e-亚基---3’?5’ 校正 ，核酸外切酶 5’?3’ exonulclease外切核酸酶在去掉引物的同时,聚合酶通过延伸临近冈崎片段3’端DNA来填补缺口. 两个片段间最后的磷酸二脂键是由DNA连接酶来催化完成的. Removal of RNA primers and filling of gaps Synthesis of Okazaki fragments requires priming, extension, removal of RNA, gap filling, and nick刻痕, 缺口ligation结扎, 绑 Termination and Segregation终止与分离 Termination site: approximately 180° opposite oriC. 复制结束时,两个子链仍是相扣的. Topoisomerase IV拓扑异构酶IV unlink the interlinked daughter genomes