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1、Chapter 8 DNA Replication Chapter 8 DNA 复制 DNA Replication Genetic information is also passed from generation to generation. A crucial property of life is that even the most complex organisms are able to be copied. When one of your cells divides by mitosis, for example, it makes two new cells that a
2、re essentially perfect replicas of the parent cell. 遗传信息也从一个世代 到另一个世代进行传递 。生命至关重要的特性 是：即使是最复杂的生 命也能够被复制出来。 例如，当你的细胞进行 有丝分裂时，它能产生 两个本质上是母细胞完 美复制品的新细胞。 basic concept DNA replication:亲代dsDNA在DNA聚合酶的作用 下，分别以ssDNA为模板，聚合与自身碱基互补配 对的游离的dNTP，合成出两条与亲代DNA完全相 同的子代DNA分子的过程。 Replicon（复制子） A unit of the genome in
3、which DNA contain a region from origin to terminator Replisome(复制体) The multiprotein(about30) structure that assembles at replicating fork to undertake synthesis of DNA. 8.1 General features of DNA replication 8.1 Semi-Conservative Replication / 半保留复制 Semi-conservative replication: A style of DNA re
4、plication in which produces a DNA with one strand from the parent, and one newly synthesized strand. 半保留复制：一种DNA的 复制方式，产生的DNA中一 条链来自于母本、另一条链 是新合成的。 8.2 Initiation of Replication / 复制的起始 OriC two critical repeated motifs: initial site: 13-mer motif, repeated three times, ssDNA formation during initia
5、tion. binding site: 9-mer motif for DnaA, repeated four times at oriC. 8.2 Initiation of Replication / 复制的起始 E. coli replication begins at a site called the OriC Replication direction New DNA enlongation from 3 5direction 磷酸基团间的强负 电性使dNTP难以 聚合，5端碱基配 对困难. Error base pairing.repair Bidirectional Repli
6、cation / 双向复制 DNA的双向复制（Replication is bidirectional ） The first issue concerns the origin of replication. Where along the chromosome is DNA replication initiated? Is there only a single origin, or does DNA synthesis begin at more than one point? Is a point of origin random or is it located at a spec
7、ific region along the chromosome? Second ,once replication begins, does it proceed in a single direction or in both directions away from the origin? In other words, is replication unidirectional or bidirectional? Replication fork : at the actual point along the chromosome where replication is occurr
8、ing, the strands of the helix are unwound, creating a replication fork. Such a fork initially appears at the point of origin of synthesis and then moves along the DNA duplex as replication proceeds. If replication is bidirectional, two such forks will be present, migrating in opposite directions awa
9、y from the origin. Cairns： Replication model（1963） 8.3 Semi-discontinuous replication Semi-discontinuous replication / 半不连续复制 The continuous strand, or leading strand, is the one in which 53synthesis proceeds in the same direction as replication fork movement. The discontinuous strand or lagging str
10、and, is the one in which 53synthesis proceeds in the direction opposite to the direction of fork movement. and, is the one Leading strand encoded by the polA gene 5 3 polymerase activity: nucleotides polymerization ,1000nt/min。 3-5 exonuclease activity :remove incorrect (mismatched) bases. 5 -3 exon
11、uclease activity: dsDNA(切除嘧啶嘧啶 二聚体) remove distorted (mispaired) segments; excision the primer. Polymerase I : three Active Sites on Its Single Polypeptide Chain Hans Klenow 用枯草杆菌蛋白酶酶（subtilisin）或 胰蛋白酶酶（trypsin）进进行降解， DNA聚合酶酶I被切 成两个片段。 smaller fragment: 5- 3exonuclease activity larger fragment :Klen
12、ow fragment has the polymerase and 3- 5exonuclease activities. DNA Polymerase 多亚基酶，分子量：120kD，每个Ecoli.细胞 100个酶分子。 催化活性： 5 3 聚合酶活性，但活力低 ,只有DNA Polymerase I 的5； 3 5 外切酶活性。 其它生理功能尚不清楚，可能在DNA的修复中起某些 作用。 DNA polymerase III / DNA聚合酶III DNA polymerase III is used for almost all of replication The full colle
13、ction of subunits is called the holoenzyme. The minimal collection of subunits needed for polymerization is called the DNA polymerase III core DNA聚合酶III 二聚体 dimer Catalytic core: Subunit-has the DNA polymerase activity. Subunit-3-5 proofreading exonuclease activity Subunit-stimulate exonuclease Its
14、two subunits clamp around the DNA template and also bind to the core, preventing it from falling off the template. Sliding clamp / 滑行夹 donut shaped protein little processivity 10 bp before detaching from the template Sliding clamp / 滑行夹 The core polymerase alone is able to polymerize DNA, but it has
15、 very little processivity. The core polymerase can only polymerize approximately 10 nucleotides before detaching from the template. The subunit(sliding clamp), is a donut shaped protein, like helicase. Its two subunits clamp around the DNA template and also bind to the core, preventing it from falli
16、ng off the template. Dimer is ring shaped, assembly or removed required energy. Dimer bound to DNA but slides along Dimer endows holoenzyme with highly processivity. Safety clamp for climbing / 攀岩安全扣 The function of sliding clamp is just like the safety clamp being used in activities like climbing. structure of sliding clamp File: E coli DNA pol III beta dimer 1MMI.val Clamp loader / 滑行夹加载器
