[预防医学]Section E-DNA复制.pptVIP

E3-1: In vitro experimental systems Purified DNA All the proteins and other factors for its complete replications DNA replication Small animal viruses (simian virus 40, 5 kb) are good mammalian models for elongation (replication fork) but not for initiation. 2. Yeast (Saccharomyces cerevisiae): 1.4 X 104 kp in 16 chromosomes, 40replicons, much simpler than mammalian system and can serve as a model system 3. Cell-free extract prepared from Xenopus (frog) eggs containing high concentration of replication proteins and can support in vitro replication. E3-2: Cell cycle When to replicate？ DNA replication G1 preparing for DNA replication (cell growth) S DNA replication G2 a short gap before mitosis M mitosis and cell division Cell cycle Entry into the S-phase: Cyclins Cyclin-dependent protein kinases (CDKs) signaling Initiaon of multiple replicons DNA replication Timing Order 大约 20-50 个复制子成簇（Clusters）在S期的同一时间开始复制 。 Early S-phase: euchromatin（常染色质） replication Late S-phase: heterochromatin（异染色质） replication Centromeric（着丝粒） and telomeric（端粒） DNA replicate last 2. Only initiate once per cell cycle Licensing factor: 起始所必须的特定蛋白（起始因子）只有在核膜解体时才能进入核内。 起始因子在作用后立即失活。 E3-3: Replication fork elongation DNA replication unwinding enzymes Replication fork Unwinding DNA from parental nucleosomes before replication : 50 bp/sec, helicases and RP-A（某种单链结合蛋白） New nucleosomes are assembled to DNA from a mixture of old and newly synthesized histones after the fork passes. Elongation: three different DNA polymerases are involved. DNA pol a: 具有引物酶的活性，在前导链的5‘端以及后随链中每个冈崎片断的5’端合成RNA引物，并继续延伸DNA链，但很快被另外的酶所代替。 DNA pol d: 在前导链上代替 DNA pol a延伸DNA链。 DNA pol e: 在后随链上代替DNA pol a合成冈崎片断。 真核生物的冈崎片断比较短(135 bp in SV40)，与核小体的结构有关。 E3-4: Telomere replication（端粒的复制） DNA replication Solving the problem of daughter DNA’s ends synthesis 5’ 3’ 5’ 3’ 3’ 3’ 5’ 5’ 3’ 5’ 3’ Parental DNA Daughter DNAs 5’ 如果染色体末端的复制问题不能解决，端粒在每个分裂过程中不断缩短，最终将导致染色体的破碎和重组。 1984年，美国加州大学的CW.Greider和EH.Blackburn