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Structural and Biochemical Characterization of the Orf9b-Tom70 Equilibrium and Opportunities to Inhibit It
Structural and Biochemical Characterization of the Orf9b-Tom70 Equilibrium and Opportunities to Inhibit It
상세정보
- 자료유형
- 학위논문 서양
- 최종처리일시
- 20260202103207
- ISBN
- 9798280730519
- DDC
- 574
- 서명/저자
- Structural and Biochemical Characterization of the Orf9b-Tom70 Equilibrium and Opportunities to Inhibit It
- 발행사항
- [Sl] : University of California, San Francisco, 2025
- 발행사항
- Ann Arbor : ProQuest Dissertations & Theses, 2025
- 형태사항
- 121 p
- 주기사항
- Source: Dissertations Abstracts International, Volume: 86-12, Section: B.
- 주기사항
- Advisor: Fraser, James;Gross, John.
- 학위논문주기
- Thesis (Ph.D.)--University of California, San Francisco, 2025.
- 초록/해제
- 요약SARS CoV 2, the causative agent responsible for the disease COVID-19 and the COVID-19 pandemic, is a member of the Betacoronavirus family with a positive-sense single-stranded RNA genome of approximately 30,000 bases in length. In addition to structural proteins and non-structural proteins (NSP's), SARS CoV 2 also encodes several accessory proteins, one of which is Orf9b. Orf9b is a 97 amino acid protein encoded through an alternative open reading frame within the nucleocapsid gene that is capable of folding into a homodimer of beta sheets, or, as a monomeric alpha helix. As a monomeric helix, Orf9b has been experimentally shown to bind to the host mitochondrial receptor Tom70 leading to the suppression of innate immunity. How Orf9b switches between these two distinct conformational and oligomeric states to bind to Tom70, as well as how this switch is regulated by the cell, is unknown.In the first chapter of this thesis, I will describe a coupled binding equilibrium between Orf9b and Tom70. I will show that Orf9b can be stabilized in a conformation and oligomeric state that is unique to the experimentally resolved structure of Orf9b bound to Tom70 upon binding of lipids to Orf9b in a homodimer conformation and use computational models to quantify the effect of lipid binding on Orf9b homodimer stability. I will also show how mutations to Orf9b observed in viral variants of concern as well as artificial Orf9b constructs alter the stability of Orfb9 in the homodimer conformation and its effects on both the equilibrium with Tom70 and innate immune activation.Following upon this work, in the second chapter, I explored the targetable space of both Orf9b and Tom70 by performing two different high-throughput screens: a crystallographic fragment screen against the Orf9b homodimer and a fluorescence polarization screen against Tom70 to identify the initial chemical matter that can be used to probe the Orf9b-Tom70 equilibrium and the mechanisms that result in innate immune suppression. I will characterize how these different small molecules can either stabilize the Orf9b homodimer to slow or inhibit binding to Tom70 or how they can bind to Tom70 to inhibit Orf9b binding. Leveraging these small molecules, I will also describe how these compounds could be used as tools to probe the Orf9b-mediated mechanism of innate immune suppression through Tom70.
- 일반주제명
- Biochemistry
- 일반주제명
- Cellular biology
- 일반주제명
- Pharmacology
- 일반주제명
- Molecular biology
- 일반주제명
- Immunology
- 기타저자
- University of California, San Francisco Biochemistry and Molecular Biology
- 기본자료저록
- Dissertations Abstracts International. 86-12B.
- 전자적 위치 및 접속
- 로그인 후 원문을 볼 수 있습니다.
MARC
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■006m o d
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■020 ▼a9798280730519
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■040 ▼aMiAaPQ▼cMiAaPQ
■0820 ▼a574
■1001 ▼aSan Felipe, Clemente.▼0(orcid)0000-0002-2695-5951
■24510▼aStructural and Biochemical Characterization of the Orf9b-Tom70 Equilibrium and Opportunities to Inhibit It
■260 ▼a[Sl]▼bUniversity of California, San Francisco▼c2025
■260 1▼aAnn Arbor▼bProQuest Dissertations & Theses▼c2025
■300 ▼a121 p
■500 ▼aSource: Dissertations Abstracts International, Volume: 86-12, Section: B.
■500 ▼aAdvisor: Fraser, James;Gross, John.
■5021 ▼aThesis (Ph.D.)--University of California, San Francisco, 2025.
■520 ▼aSARS CoV 2, the causative agent responsible for the disease COVID-19 and the COVID-19 pandemic, is a member of the Betacoronavirus family with a positive-sense single-stranded RNA genome of approximately 30,000 bases in length. In addition to structural proteins and non-structural proteins (NSP's), SARS CoV 2 also encodes several accessory proteins, one of which is Orf9b. Orf9b is a 97 amino acid protein encoded through an alternative open reading frame within the nucleocapsid gene that is capable of folding into a homodimer of beta sheets, or, as a monomeric alpha helix. As a monomeric helix, Orf9b has been experimentally shown to bind to the host mitochondrial receptor Tom70 leading to the suppression of innate immunity. How Orf9b switches between these two distinct conformational and oligomeric states to bind to Tom70, as well as how this switch is regulated by the cell, is unknown.In the first chapter of this thesis, I will describe a coupled binding equilibrium between Orf9b and Tom70. I will show that Orf9b can be stabilized in a conformation and oligomeric state that is unique to the experimentally resolved structure of Orf9b bound to Tom70 upon binding of lipids to Orf9b in a homodimer conformation and use computational models to quantify the effect of lipid binding on Orf9b homodimer stability. I will also show how mutations to Orf9b observed in viral variants of concern as well as artificial Orf9b constructs alter the stability of Orfb9 in the homodimer conformation and its effects on both the equilibrium with Tom70 and innate immune activation.Following upon this work, in the second chapter, I explored the targetable space of both Orf9b and Tom70 by performing two different high-throughput screens: a crystallographic fragment screen against the Orf9b homodimer and a fluorescence polarization screen against Tom70 to identify the initial chemical matter that can be used to probe the Orf9b-Tom70 equilibrium and the mechanisms that result in innate immune suppression. I will characterize how these different small molecules can either stabilize the Orf9b homodimer to slow or inhibit binding to Tom70 or how they can bind to Tom70 to inhibit Orf9b binding. Leveraging these small molecules, I will also describe how these compounds could be used as tools to probe the Orf9b-mediated mechanism of innate immune suppression through Tom70.
■590 ▼aSchool code: 0034.
■650 4▼aBiochemistry
■650 4▼aCellular biology
■650 4▼aPharmacology
■650 4▼aMolecular biology
■650 4▼aImmunology
■653 ▼aCoupled equilibrium
■653 ▼aProtein biochemistry
■653 ▼aSmall molecule discovery
■653 ▼aStructural biology
■653 ▼aCOVID-19 pandemic
■690 ▼a0487
■690 ▼a0379
■690 ▼a0982
■690 ▼a0419
■690 ▼a0307
■71020▼aUniversity of California, San Francisco▼bBiochemistry and Molecular Biology.
■7730 ▼tDissertations Abstracts International▼g86-12B.
■790 ▼a0034
■791 ▼aPh.D.
■792 ▼a2025
■793 ▼aEnglish
■85640▼uhttp://www.riss.kr/pdu/ddodLink.do?id=T17357319▼nKERIS▼z이 자료의 원문은 한국교육학술정보원에서 제공합니다.
