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Evolutionary Motif Swapping of Human Dihydrofolate Reductase Rewires the Enzymatic Cycle
Evolutionary Motif Swapping of Human Dihydrofolate Reductase Rewires the Enzymatic Cycle
상세정보
- 자료유형
- 학위논문 서양
- 최종처리일시
- 20260202103557
- ISBN
- 9798280710382
- DDC
- 574
- 서명/저자
- Evolutionary Motif Swapping of Human Dihydrofolate Reductase Rewires the Enzymatic Cycle
- 발행사항
- [Sl] : Harvard University, 2025
- 발행사항
- Ann Arbor : ProQuest Dissertations & Theses, 2025
- 형태사항
- 111 p
- 주기사항
- Source: Dissertations Abstracts International, Volume: 86-12, Section: B.
- 주기사항
- Advisor: Hekstra, Doeke R.
- 학위논문주기
- Thesis (Ph.D.)--Harvard University, 2025.
- 초록/해제
- 요약Despite common descent, enzymes often rescue poorly when expressed across domains of life. Causes include factors external to the enzyme, such as differences in codon usage, sensitivity to proteases, and transcriptional or post-translational regulatory differences, yet often the underlying cause remains unclear. Dihydrofolate reductase (DHFR) catalyzes the same metabolic conversion across the tree of life. Nevertheless, human DHFR (hsDHFR) does not effectively rescue growth of DHFR-deficient E. coli despite similar in vitro kinetics. This phenomenon has been previously attributed to inhibition of hsDHFR by its oxidized cofactor, NADP+. To understand this phenomenon, we designed mutants based on deep sequence divergences across the tree of life, yielding variants which outperform both wild-type enzymes in vitro and which rescue growth of E. coli. Remarkably, a single, ancient sequence insertion underlies gain of function, not by modulating product inhibition, but by redirecting ligand flux - the non-equilibrium sequence of steps binding and unbinding product, cofactor, and substrate. We find that deleting this insertion decouples the dynamics of a substrate binding loop from subdomain motion, thereby controlling a critical enzymatic parameter orthogonal to catalytic proficiency.
- 일반주제명
- Biochemistry
- 일반주제명
- Biophysics
- 일반주제명
- Biology
- 일반주제명
- Microbiology
- 키워드
- E. coli
- 기타저자
- Harvard University Biology Molecular and Cellular
- 기본자료저록
- Dissertations Abstracts International. 86-12B.
- 전자적 위치 및 접속
- 로그인 후 원문을 볼 수 있습니다.
MARC
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■040 ▼aMiAaPQ▼cMiAaPQ
■0820 ▼a574
■1001 ▼aBrookner, Dennis E.▼0(orcid)0000-0003-2827-4743
■24510▼aEvolutionary Motif Swapping of Human Dihydrofolate Reductase Rewires the Enzymatic Cycle
■260 ▼a[Sl]▼bHarvard University▼c2025
■260 1▼aAnn Arbor▼bProQuest Dissertations & Theses▼c2025
■300 ▼a111 p
■500 ▼aSource: Dissertations Abstracts International, Volume: 86-12, Section: B.
■500 ▼aAdvisor: Hekstra, Doeke R.
■5021 ▼aThesis (Ph.D.)--Harvard University, 2025.
■520 ▼aDespite common descent, enzymes often rescue poorly when expressed across domains of life. Causes include factors external to the enzyme, such as differences in codon usage, sensitivity to proteases, and transcriptional or post-translational regulatory differences, yet often the underlying cause remains unclear. Dihydrofolate reductase (DHFR) catalyzes the same metabolic conversion across the tree of life. Nevertheless, human DHFR (hsDHFR) does not effectively rescue growth of DHFR-deficient E. coli despite similar in vitro kinetics. This phenomenon has been previously attributed to inhibition of hsDHFR by its oxidized cofactor, NADP+. To understand this phenomenon, we designed mutants based on deep sequence divergences across the tree of life, yielding variants which outperform both wild-type enzymes in vitro and which rescue growth of E. coli. Remarkably, a single, ancient sequence insertion underlies gain of function, not by modulating product inhibition, but by redirecting ligand flux - the non-equilibrium sequence of steps binding and unbinding product, cofactor, and substrate. We find that deleting this insertion decouples the dynamics of a substrate binding loop from subdomain motion, thereby controlling a critical enzymatic parameter orthogonal to catalytic proficiency.
■590 ▼aSchool code: 0084.
■650 4▼aBiochemistry
■650 4▼aBiophysics
■650 4▼aBiology
■650 4▼aMicrobiology
■653 ▼aProtein structure
■653 ▼aStructural biology
■653 ▼aX-ray crystallography
■653 ▼aE. coli
■653 ▼aDihydrofolate reductase
■690 ▼a0487
■690 ▼a0786
■690 ▼a0306
■690 ▼a0410
■71020▼aHarvard University▼bBiology, Molecular and Cellular.
■7730 ▼tDissertations Abstracts International▼g86-12B.
■790 ▼a0084
■791 ▼aPh.D.
■792 ▼a2025
■793 ▼aEnglish
■85640▼uhttp://www.riss.kr/pdu/ddodLink.do?id=T17357767▼nKERIS▼z이 자료의 원문은 한국교육학술정보원에서 제공합니다.
