자료유형  

 학위논문

Control Number  

0017162468

International Standard Book Number  

9798382837314

Dewey Decimal Classification Number  

574

Main Entry-Personal Name  

Gonzalez-DeWhitt, Kristofer R.

Publication, Distribution, etc. (Imprint  

[S.l.] : University of California, Los Angeles., 2024

Publication, Distribution, etc. (Imprint  

Ann Arbor : ProQuest Dissertations & Theses, 2024

Physical Description  

297 p.

General Note  

Source: Dissertations Abstracts International, Volume: 85-12, Section: B.

General Note  

Advisor: Abramson, Jeff S.

Dissertation Note  

Thesis (Ph.D.)--University of California, Los Angeles, 2024.

Summary, Etc.  

요약Cannabinoids are produced in Cannabis sativa L. via a highly modular process involving chemical precursors originating from the polyketide synthase and monoterpene biosynthetic pathways. In the polyketide synthase pathway, tetraketide synthase (TKS) catalyzes an imprecise reaction to produce a diffusible, linear tetra-β-ketide that is subsequently cyclized by an accessory protein to olivetolic acid. Alkylation, oxidative cyclization, and non-enzymatic decarboxylation of molecules derived from olivetolic acid produce the pharmaceutically important cannabinoids (-)-trans-Δ9 - tetrahydrocannabinol (Δ9 -THC) and cannabidiol (CBD). Besides producing a required pathway precursor, TKS generates multiple aberrant by-products that derail cannabinoid biosynthesis. Structure determination and site-directed mutagenesis experiments were conducted to clarify the TKS reaction mechanism. Crystal structures of TKS in complex with coenzyme A and a pentyl mono-β-ketide, pentyl di-β-ketide, or pentyl tri-β-ketide revealed that polyketide synthesis is facilitated by horizontal expansion of the cyclization pocket. Horizontal expansion is mediated by a single residue rotating outwards from the cyclization pocket. Substitutions truncating this residue's sidechain improved olivetolic acid formation by 176%. Orthogonal structure-function experiments were conducted on hexaketide synthase (Drosophyllum lusitanicum) (DluHKS) and implicated a structural basis for aberrant by-products catalyzed by TKS. A crystal structure was determined for DluHKS and revealed a pentyl tri-β-ketide covalently attached in monomer A and a covalent pentyl di-β-ketide in monomer B. The pentyl tri-β-ketide is stabilized within the cyclization pocket via a hydrogen bond that additionally predisposes polyketide intermediates to lactonization. These collective efforts to determine the structural basis of TKS and DluHKS catalytic activity provide an updated understanding of the TKS reaction mechanism as well as suggestions for rationally mutating TKS for improved cannabinoid bioproduction.

Subject Added Entry-Topical Term  

Biochemistry.

Subject Added Entry-Topical Term  

Biology.

Subject Added Entry-Topical Term  

Molecular biology.

Subject Added Entry-Topical Term  

Bioengineering.

Index Term-Uncontrolled  

Cannabinoids

Index Term-Uncontrolled  

Crystallography

Index Term-Uncontrolled  

Olivetolic acid

Index Term-Uncontrolled  

Structural biology

Index Term-Uncontrolled  

Tetraketide synthase

Added Entry-Corporate Name  

University of California, Los Angeles Molecular Biology 0573

Host Item Entry  

Dissertations Abstracts International. 85-12B.

Electronic Location and Access  

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Control Number  

joongbu:654671