Title

Characterization of Endospore Proteins Associated with DNA Protection

Alternative Title

디옥시리보핵산 보호와 관련된 내생포자 단백질의 특성 분석

DGIST Authors

Hyogyung Shin ; Yoori Kim ; Young-Sam Lee

Advisor

김유리

Co-Advisor(s)

Young-Sam Lee

Issued Date

2023

Awarded Date

2023-08-01

Citation

Hyogyung Shin. (2023). Characterization of Endospore Proteins Associated with DNA Protection. doi: 10.22677/THESIS.200000686904

Type

Thesis

Description

Clostridium perfringens; endospore; small acid-soluble protein; DNA protection; sortase-mediated fluorescent labeling

Table Of Contents

Ⅰ. Introduction 1
Ⅱ. Materials and Methods 9
2.1 Wild-type Ssp4 expression plasmids 9
2.2 Purification of Ssp4 9
2.3 Preparation of DNA 10
2.4 Electrophoretic mobility shift assay (EMSA) 11
2.5 DNA protection assay against UV irradiation 12
2.6 DNA protection assay against DNase I degradation 13
2.7 Site-directed mutagenesis for N-terminal GGG-Ssp4 14
2.8 Purification of sortase A 15
Ⅲ. Results 17
3.1 Purification of Ssp4:D36 and Ssp4:G36 proteins 17
3.2 DNA binding affinity for Ssp4 variants for random-sequence DNA 20
3.3 DNA binding affinity for Ssp4 variants for AT-rich DNA 24
3.4 DNA protection activity of Ssp4 variants 27
3.5 Sortase-mediated fluorescent labeling 31
3.5.1 Construction of N-terminal GGG-Ssp4 vector 31
3.5.2 Purification of sortase A 34
3.5.3 Establishment of sortase-mediated fluorescent labeling strategy for Ssp4 36
Ⅳ. Discussion 40
References 44
Summary in Korean 56

URI

http://hdl.handle.net/20.500.11750/46423
http://dgist.dcollection.net/common/orgView/200000686904

DOI

10.22677/THESIS.200000686904

Degree

Master

Department

Department of New Biology

Publisher

DGIST

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