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1、参与原核生物DNA复制的酶类和蛋白质Enzymes and Proteins Involved in DNA Replication in Prokaryotes 高方远 马欣荣 康海岐,DNA replication(bacteria)InitiationElongationTerminationDaughter DNA partition,*the origin of replication is defined*replication bubble.*Replication fork*Unidirectional or bidirectional.,Origins,Elongation(
2、semidiscontinuous),termination,参与DNA复制的酶类,1、DNA聚合酶2、DNA引发酶（DNA primase）3、DNA连接酶4、与DNA几何学性质相关的酶,I polA major repair enzyme II polB minor repair enzyme III polC replicase IV dinB SOS repair V umuC、D SOS repair,DNA polymerases in E.coli,DNA Polymerase I,DNA polymerase3 5 exonuclease5 3 exonuclease,DNA
3、Polymerase I,Subunit composition of E.coli DNA polymerase III holoenzyme,E.coli Pol III Beta-subunit,Fig.1.Model of SOS translesion replication by DNA polymerase V.The two DNA strands are shown as green lines,and the replication-blocking lesion is represented by the red rectangle.The three major ste
4、ps in TLR are pre-initiation(2),in which the RecA nucleoprotein filaments assembles;initiation(3and 4),which involves binding of pol V to the primer-template and loading of the subunit clamp;and lesion bypass by pol V holoenzyme(5).SSB is suggested to help in displacing RecA from DNA both at the ini
5、tiation and lesion bypass steps.,2、DNA引发酶（DNA primase）,Use host RNA polymerase as primase(M13)primosome primase(dnaG protein)(E.coli)other proteins X174:only primase,without the other proteins,DNA Polymerase I,与DNA几何学性质相关的酶,解旋酶（Helicase）拓扑异构酶（Topoisomerases）,解旋酶（Helicase）,至少4种helicases*rep helicase*
6、DNA helicase II*DNA helicase III*dnaB Protein:在E.coli DNA复制中解开 DNA双链,拓扑异构酶（Topoisomerases）,拓扑异构酶I(topA gene)act on highly negatively supercoiled DNA stabilize single-stranded regions,Figure 14.16 Bacterial type I topoisomerases recognize partially unwound segments of DNA and pass one strand through
7、a break made in the other.,拓扑异构酶II,Type II topoisomerases generally relax both negative and positive supercoils.The reaction requires ATP,Figure 14.17 Type II topoisomerases can pass a duplex DNA through a double-strand break in another duplex.,拓扑异构酶IV,与子代DNA分子的分开有关,参与DNA复制的蛋白质,1、参与复制起始的蛋白质因子Origina
8、l complx:DnaA、DnaB、DnaC、DnaG、HUand SSB,SeqA,Antibodies of dnaA,protein HU,The protein HU is a general DNA-binding protein in E.coli.Its presence is not absolutely required to initiate replication in vitro,but it stimulates the reaction.HU has the capacity to bend DNA,and is likely to be involved in
9、some general structural capacity.,2、参与复制延伸的蛋白质因子,(Dna G),(DnaB),2、参与复制终止的蛋白质因子,Tus,How do the daughter DNAs become disentangled?,与真核生物不同，细菌的DNA复制、染色体重新折叠以及姊妹染色体的分开是同时发生的。细菌中姊妹染色体的分开的机制与真核生物不同。细菌染色体的DNA分子本身卷入了分开机制。细菌的多复制叉复制（multiple replication）与真核生物的复制方式不同。*多拷贝的oriC*只有一个终止序列,SMC类似物,复制起点及终点在细胞中的位置,1、
10、膜片段中有复制起始区与终止区的富集推断锚定蛋白在定位中的作用。,SeqA,2、位于中间位置 复制工厂 的动力作用 多蛋白复合体：polymerase,helicase and accociated proteins 特殊蛋白质（PolC-GFP、SeqA)的定位；H3同位素标记；pull DNA template duplicated release DNA outward during replication,The extrusion-capture model for bacterial chromosome partitioning.,3、与染色体组织（organization）、紧结
11、(compaction)和超螺旋(helicase)有关的蛋白质作用 SMC MukB to organize the chromosome into a higher order structure by constraining supercoils.(cause),Partitioning Motor protein(altered),Chromosome partitioning(consequence),HU;Hbsu;,Tyrosine site-specific recombinases,E.coli,CodV,RipXB.subtilis,Post-septationpartitioning,FtsK,PBP2 RodA,PBP 3 peptidoglycan(肽聚糖),EnvA,Minicells:anucleate cells,Problem哪些DNA位点和蛋白质对复制起点的移动及定位起决定性作用？这些位点和蛋白质在指数生长期和进入静止期是否相同？细胞分裂后，终止序列是怎样定位到细胞的中央部位的？能够进行多复制叉复制的细菌与不能进行多复制叉复制的细菌在染色体分开机制上有什么不同？不同物理形状的染色体分开时有差异吗？,？,Thank you!,20.,
