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Dissertation
Dirigent protein structure and function
Washington State University
Doctor of Philosophy (PhD), Washington State University
2023
DOI:
https://doi.org/10.7273/000004986
Abstract
This PhD thesis focuses on dirigent protein (DP) functions and biochemical mechanisms in two nitrogen-fixing (N-fixing) organisms, pea (Pisum sativum) and red alder (Alnus rubra Bong). There are two main objectives: 1) Investigate DP functions, biochemical mechanisms, and DP 3D structures in pea and licorice; and 2) Assess DP sub-families in red alder through genome sequencing and integrated RNA-Seq/metabolomics analyses. Objective 1: This objective studied 3 licorice and pea DPs which produce antimicrobials. Each DP had kinetic parameters and 3-D X-ray crystal structures determined. Stable isotope labelling of the licorice DP and its NMR analyses explored how it and its mutants dynamically interact with substrates. An integrated bottom-up, top-down, native mass spectrometry approach was also deployed to de novo investigate potential Forsythia DP interacting proteins.
Objective 2: This focused on red alder and its possible DP sub-family biochemical functions. Red alder (clone 639) was sequenced, with 52,758 coded protein genes being predicted and annotated including 29 DPs in 3 sub-families. Clone 639 was subjected to metabolomics and RNA-seq analyses at three different time points in the growing season. Targeted and non-targeted metabolomics established ~657 features with 72 metabolites either identified, annotated, or provisionally annotated. Together with RNA-seq data, our bioinformatics analysis focused upon red alder’s beneficial plant defenses, its reddish-orange dye pigments, and its medicinal compounds, with particular attention given to the hypothesized DP gateway entry points to the bioactive proanthocyanidins, ellagitannins and diarylheptanoids giving red alder such positive attributes.
The PhD thesis has nine chapters: Chapters 2, 4, 5, 7 and 8 are published in Journal of Biological Chemistry, Methods in Enzymology, Analyst, G3-Genes Genomes Genetics, and The Lignan Handbook, respectively. Chapters 3, 6 and 9 are in finalization and will be submitted for publication shortly. Chapter 1 is a brief introduction to the topic area and justification for the research carried out.
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Details
- Title
- Dirigent protein structure and function
- Creators
- Qingyan Meng
- Contributors
- Norman G. Lewis (Advisor)John R. Cort (Committee Member)Michael L. Kahn (Committee Member)Michael Knoblauch (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- Program in Molecular Plant Sciences
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Publisher
- Washington State University
- Number of pages
- 476
- Identifiers
- 99901019834701842
- Language
- English
- Resource Type
- Dissertation