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首页> 外文学位 >THE INTERACTION OF BASE-ANALOG SUBSTITUTED LAMBDA P(,R) PROMOTERS WITH ESCHERICHIA COLI RNA POLYMERASE (DNA SYNTHESIS).
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THE INTERACTION OF BASE-ANALOG SUBSTITUTED LAMBDA P(,R) PROMOTERS WITH ESCHERICHIA COLI RNA POLYMERASE (DNA SYNTHESIS).

机译：碱基类似物取代的Lambda P（，R）启动子与大肠埃希氏菌RNA聚合酶的相互作用（DNA合成）。

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摘要

Transcription in E. coli is catalyzed by DNA dependent RNA polymerase. This enzyme specifically recognizes the promoter, the DNA sequence at which transcription is initiated. Two regions of highly conserved sequence are located approximately 33 and 10 base pairs away from the site of transcription initiation (+1). These sequences are TTGACA near -35 and TATAAT near -10. A promoter for E. coli RNA polymerase, the rightward promoter of bacteriophage lambda ((lamda)P(,R)), was synthesized by a combination of chemical and enzymatic methods. Site-specific base changes were introduced into the synthetic promoter at regions thought to be important in the specific recognition of E. coli promoters by polymerase. Most base modifications were found to have negligible effect on the activity of the promoter. A few changes at positions of highly conserved sequence in E. coli promoters were found to reduce promoter activity severely or to yield promoters of intermediate strength.Two methyl groups in the -35 region appear to have a composite effect on the function of the promoter. Removal of both of them inactivates the promoter, while removal of either one creates a promoter showing slower reaction kinetics with RNA polymerase. Interaction of polymerase with a strongly conserved T(.)A base pair in the last position of the -10 homology seems to be taking place in the major groove. These results are discussed in relation to current knowledge of E. coli RNA polymerase function.Removal of both methyl groups from the thymidines at the first two positions of the -35 homology by introduction of two uridines inactivated the promoter. This result was further investigated by making the corresponding single modifications in the -35 region. The two promoters having the sequence TUGACA or UTGACA in the -35 homology were functional but displayed slower reaction kinetics than the unmodified promoter. In the -10 region replacement of the T(.)A base pair at -7 with a C(.)I base pair also inactivated the promoter. The only difference between the C(.)I and the wild type T(.)A base pair is found in the major groove. This result suggests that interaction with RNA polymerase at this position occurs primarily in the major groove.
机译：DNA依赖性RNA聚合酶可催化大肠杆菌中的转录。该酶特异性识别启动子，即开始转录的DNA序列。高度保守序列的两个区域位于距转录起始位点（+1）大约33和10个碱基对的位置。这些序列是-35附近的TTGACA和-10附近的TATAAT。大肠杆菌RNA聚合酶的启动子，即噬菌体λ（λP（，R））的向右启动子，是通过化学和酶促方法相结合而合成的。将位点特异性碱基改变引入合成启动子中被认为对聚合酶特异性识别大肠杆菌启动子重要的区域。发现大多数碱基修饰对启动子的活性影响可忽略。发现在大肠杆菌启动子中高度保守的序列位置上的一些变化会严重降低启动子活性或产生中等强度的启动子。-35区中的两个甲基似乎对启动子的功能具有复合作用。去除它们两者都使启动子失活，而去除其中任何一个都会产生显示与RNA聚合酶反应动力学较慢的启动子。聚合酶与强保守的T（。）A碱基对在-10同源性的最后一个位置的相互作用似乎发生在主要沟中。这些结果是关于大肠杆菌RNA聚合酶功能的当前知识而讨论的。通过引入两个尿苷使启动子失活，从-35同源性的前两个位置上的胸苷中去除两个甲基。通过在-35区域进行相应的单个修饰，可以进一步研究该结果。在-35同源性中具有序列TUGACA或UTGACA的两个启动子是功能性的，但是显示出比未修饰的启动子慢的反应动力学。在-10区，用C（.I）碱基对替换在-7处的T（。）A碱基对也使启动子失活。 C（。）I和野生型T（。）A碱基对之间的唯一区别是在主沟中。该结果表明在该位置与RNA聚合酶的相互作用主要发生在主要沟中。

著录项

  • 作者

    DUBENDORFF, JOHN WOODS.;

  • 作者单位

    University of Colorado at Boulder.;

  • 授予单位 University of Colorado at Boulder.;
  • 学科 Chemistry Biochemistry.
  • 学位 Ph.D.
  • 年度 1985
  • 页码 144 p.
  • 总页数 144
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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