MARC 主機 00000nam a2200541K 4500
001 AAI27726326
005 20201105110322.5
006 m o d
007 cr mn ---uuuuu
008 201105s2019 miu sbm 000 0 eng d
020 9781392878248
035 (MiAaPQ)AAI27726326
035 (MiAaPQ)0098vireo4626Bettridge
040 MiAaPQ|beng|cMiAaPQ|dNTU
100 1 Bettridge, Kelsey Elaine
245 10 Probing The Dynamics of RNA Polymerase and Hu in Live E.
coli Cells
264 0 |c2019
300 1 online resource (185 pages)
336 text|btxt|2rdacontent
337 computer|bc|2rdamedia
338 online resource|bcr|2rdacarrier
500 Source: Dissertations Abstracts International, Volume: 81-
08, Section: B
500 Advisor: Wolberger, Cynthia
502 Thesis (Ph.D.)--The Johns Hopkins University, 2019
504 Includes bibliographical references
520 Diffusion is at the heart of every biochemical process.
Millions of proteins must navigate the heterogeneous,
crowded cellular milieu to perform their various tasks.
This molecular crowding has a significant effect on the
diffusive behavior and kinetic rates of proteins and
biochemical reactions. A powerful technique to understand
biochemical processes within the context of this
heterogeneous environment is single particle tracking
(SPT). In Chapter 2, I use SPT to elucidate the dynamics
of the RNAP search process and transcription cycle in live
E. coli cells. Using FRAP, I find that transcription
follows a simple initiation-elongation-termination cycle
with kinetic rates that closely match those in the
literature. Using SPT, I probed the search process of RNAP
and found three diffusive states corresponding to DNA-
bound, diffusion within the dense nucleoid, and diffusion
within the cytoplasm. RNAP exhibited confinement in each
state and displayed a preference for a DNA-bound state,
suggesting a grid search strategy. Additionally, RNAP
displayed kinetics that were not consistent with steady
state kinetics. In Chapter 3, I use SPT to probe the
molecular mechanism of HU-mediated chromosome
organization. Using genetic mutations that abolish the
various binding modes of HU, I find that HUαα and HUαβ
displayed differential dynamics. Additionally, HUαα seems
primarily responsible for non-specific binding while HUαβ
seems primarily responsible for repressor loop formation.
The kinetics of HU were highly transient, indicative of
their non-specific binding across the nucleoid, and
suggested a mechanism by which cumulative forces of
thousands of HU are able to achieve chromosomal
organization, a marked departure from the long-lived
binding of other DNA organization proteins such as
histones
533 Electronic reproduction.|bAnn Arbor, Mich. :|cProQuest,
|d2020
538 Mode of access: World Wide Web
650 4 Microbiology
650 4 Optics
650 4 Biophysics
653 Biophysics
653 Single particle tracking
653 Diffusion
653 Fluorescence microscopy
653 Gene regulation
653 Chromosome organization
653 Nucleoid organization
653 RNA polymerase
653 DNA-binding protein
655 7 Electronic books.|2local
690 0786
690 0410
690 0752
710 2 ProQuest Information and Learning Co
710 2 The Johns Hopkins University.|bProgram in Molecular
Biophysics
773 0 |tDissertations Abstracts International|g81-08B
856 40 |uhttp://pqdd.sinica.edu.tw/twdaoapp/servlet/
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