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Structure of Escherichia coli H-NS and Mixed H-NS Filaments and Their Effect on Transcription Elongation by RNA Polymerase
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| 자료유형 | 학위논문 |
|---|---|
| 서명/저자사항 | Structure of Escherichia coli H-NS and Mixed H-NS Filaments and Their Effect on Transcription Elongation by RNA Polymerase. |
| 개인저자 | Shen, Beth Ann. |
| 단체저자명 | The University of Wisconsin - Madison. Biochemistry-ALS. |
| 발행사항 | [S.l.]: The University of Wisconsin - Madison., 2019. |
| 발행사항 | Ann Arbor: ProQuest Dissertations & Theses, 2019. |
| 형태사항 | 444 p. |
| 기본자료 저록 | Dissertations Abstracts International 81-04B. Dissertation Abstract International |
| ISBN | 9781687904515 |
| 학위논문주기 | Thesis (Ph.D.)--The University of Wisconsin - Madison, 2019. |
| 일반주기 |
Source: Dissertations Abstracts International, Volume: 81-04, Section: B.
Advisor: Landick, Robert. |
| 이용제한사항 | This item must not be sold to any third party vendors.This item must not be added to any third party search indexes. |
| 요약 | An assortment of bacterial chromatin proteins regulate transcription by RNA polymerase (RNAP) through multiple mechanisms. These mechanisms are incompletely characterized because of limited structural details of chromatin proteins. In my thesis work, I studied one group of chromatin proteins (H-NS, StpA, and Hha) that silence genes in many Gram-negative bacteria, including Escherichia coli, by forming filaments on AT-rich DNA. H-NS and StpA form filaments by first binding high-affinity sites via a C-terminal DNA-binding domain (DBD) and then oligomerizing along the DNA via protein-protein interactions in an N-terminal domain. H-NS can form both a bridged filament, where H-NS interacts with two segments of DNA and a linear filament, where H-NS interacts with one segment of DNA. StpA only bridges DNA. In vivo, H-NS and StpA likely exist as heterodimers. Hha interacts with H-NS and StpA, but it does not bind to DNA. The filaments that exist in vivo are likely composed of these three proteins, but the structure and function of these mixed filaments is unknown. In the first part of my thesis work, I determined if addition of StpA or Hha to an H-NS filament modified the effect of H-NS on an elongating RNAP. Bridged, but not linear, H-NS filaments were previously shown to stimulate pausing in an elongating RNAP in vitro, and I found that both StpA and Hha stimulated bridging by H-NS, which enhanced pausing by RNAP. These results suggest that bridged mixed filaments might aid gene silencing in vivo. In the second part of my thesis work, I investigated two aspects of filament structure to better understand the mechanism of gene silencing. First, using high-throughput sequencing strategies, I found that sequence-specific binding alone dictated filament formation throughout the genome. Second, I determined that the H-NS DBDs bind in the same pattern in both bridged and linear filaments using a tethered cleavage assay. These results are consistent with a model where sequence dictates location of filament formation, but conformational changes in H-NS facilitate modes of binding. Overall, my thesis work provides insight into the factors influencing filament formation and suggests important gene regulatory roles for different filament conformations. |
| 일반주제명 | Biochemistry. |
| 언어 | 영어 |
| 바로가기 | 
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