大肠杆菌topA-菌株中质粒pBR322的超凝集结构  

Studies on The Supercondensed pBR322 DNA Isolated From Escherichia coli topA-Mutant

作　　者：张臻峰[1] 于佳[1] 曹扣[1] 黄熙泰[1] 

机构地区：[1]南开大学生命科学学院生物化学与分子生物学系,天津300071

出　　处：《生物化学与生物物理进展》2007年第3期306-311,共6页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金资助项目(39970014)~~

摘　　要：超凝集DNA是质粒DNA的一种特殊拓扑结构形式,最初在大肠杆菌SD108(topA+gyrB225)细胞中被发现.现在大肠杆菌DM800(topA-gyrB225)细胞中也发现了这种结构,这说明超凝集DNA的形成与细胞内旋转酶活性降低有直接关系,而与拓扑异构酶Ⅰ的存在与否无关.体外实验的结果显示,具有很强的正超螺旋松弛活性的拓扑异构酶Ⅳ可以将超凝集DNA完全松弛,这也证明质粒DNA的超凝集结构与超螺旋结构在细胞内是可以互变的.使用原子力显微镜对分离到的pBR322DNA超凝集结构进行分析,并与普通超螺旋进行比较,结果表明,超凝集DNA分子的结构发生了巨大的变化,其分子长度比正常超螺旋分子缩短了约30%,宽度和高度则增加了60%,结构更接近于A型DNA.另外,原子力显微镜研究结果表明,氯喹的嵌入并非改变了超凝集DNA的超螺旋状态,而是使其打结并最终压缩成一团.The supercondensed DNA, a special kind of topological structure ofplasmid DNA, was firstly found in E. coli SD108（topA＋ gyrB225）. Now, this structure is also found in E. coli DM800（topA- gyrB225）. The result indicates that the formation of supercondensed DNA is related with decrease of the activity of gyrase in vivo. Topoisomerase IV was proved to relax the supercondensed DNA completely in vitro, which suggested that the supercondensed DNA and the supercoiled DNA could transform to each other in cells. The supercondensed DNA samples were analyzed by atomic force microscopy and compared to supercoiled DNA. The results showed that the length of supercondensed DNA decreased about 30% and the width and height of double-strand increased about 60%, which indicates that the structure of double-strand of supercondensed DNA is much more similar to A-DNA than B-DNA. The results also showed that chloroquine intercalation did not change the supercoiling level of supercondensed DNA, but made it knot and compact.

关 键 词：超凝集DNA 大肠杆菌DM800 原子力显微镜 拓扑异构酶Ⅳ 

分 类 号：Q71[生物学—分子生物学]