THESIS
2020
xv, 104 pages : color illustrations ; 30 cm
Abstract
The central dogma of molecular biology, which involves replication, transcription and
translation, describes the flow of gene expression from DNA to RNA to protein. DNA converts
genetic information by transcribing RNA through RNA polymerase (RNAP) and then
translating protein by ribosomes. To maintain both efficiency and fidelity during the
transmission of genetic information to proteins, many enzymes are involved. In this thesis, the
working mechanism of two enzymes will be discussed: DNA glycosylase and RNAP. DNA
glycosylase excises lesions from DNA to maintain the integrity of the genome and RNAP
transcribes from DNA into RNA.
Alkylpurine glycosylase D (AlkD), a DNA glycosylase, is responsible for repairing DNA
damage to maintain the integrity of the genome. However, how gly...[
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The central dogma of molecular biology, which involves replication, transcription and
translation, describes the flow of gene expression from DNA to RNA to protein. DNA converts
genetic information by transcribing RNA through RNA polymerase (RNAP) and then
translating protein by ribosomes. To maintain both efficiency and fidelity during the
transmission of genetic information to proteins, many enzymes are involved. In this thesis, the
working mechanism of two enzymes will be discussed: DNA glycosylase and RNAP. DNA
glycosylase excises lesions from DNA to maintain the integrity of the genome and RNAP
transcribes from DNA into RNA.
Alkylpurine glycosylase D (AlkD), a DNA glycosylase, is responsible for repairing DNA
damage to maintain the integrity of the genome. However, how glycosylases efficiently and
accurately recognize DNA lesions among numerous base pairs has not been elucidated. It has
been hypothesized that glycosylase translocation occurs by alternating between a high speed-low
accuracy diffusion mode and a low speed-high accuracy mode in searching for lesions. A
slow mode exists when AlkD approaches lesions and is thus important for lesion searching.
However, the molecular mechanism for the slow mode has not been determined. In this study,
using the Markov state model built from extensive all-atom Molecular dynamics (MD)
simulations, we elucidated the mechanism governing the slow mode. We found that in the slow
mode, AlkD follows an asymmetric diffusion pathway, i.e., rotation followed by translation.
In addition, essential roles of Y27 and Q38 in diffusion were identified.
RNAP facilitates DNA transcription of RNA. Transcription includes initiation, elongation and
termination. Previous studies have provided crystal structures for intermediate states in the
initiation process. However, due to the large conformational changes exhibited by the RNAP-DNA
complex, the kinetic mechanism of this complex during initiation has not been elucidated.
In this study, coarse-grained MD was applied to study the initiation of the RNAP-DNA
complex. Our results suggest that the flexibility of critical regions, including fork loop 2 and
sigma fingers, gave space to the template DNA entering the active site.
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