UC Santa Cruz

Natural products have long served as a rich source of biologically active molecules and continue to drive discoveries in chemistry, biology, and medicine. Despite advances in microbial cultivation, genome mining, and synthetic biology, the pace of natural product research remains constrained by analytical workflows required to characterize chemical signaling, identify novel compounds, and evaluate engineered biosynthetic systems. Recent developments in mass spectrometry (MS), including multidimensional separations and high-throughput acquisition approaches, provide new opportunities to accelerate these workflows. This dissertation explores the development and application of high-throughput, multidimensional MS approaches to advance natural product discovery, chemical ecology, and enzyme engineering.Chapter 2 expands the capabilities of microbial MALDI-MS by integrating tandem mass spectrometry (MALDI-MS/MS) into the bioinformatics platform IDBac, enabling streamlined characterization of microbial taxonomy, metabolite production, and preliminary metabolite annotation directly from microbial colonies. Application of this workflow enabled the annotation of diverse microbial natural products, including lavanducyanin, napyradiomycins, surugamides, antimycins, and desferrioxamine siderophores, culminating in the discovery of napyradiomycin B8 and the characterization of harmane derivatives.Chapter 3 advances microbial mass spectrometry imaging (MSI) by combining MALDI imaging with trapped ion mobility spectrometry (TIMS) and imaging parallel reaction monitoring-parallel accumulation serial fragmentation (iprm-PASEF). This workflow improves metabolite annotation by incorporating targeted MS/MS acquisition directly into MALDI-MSI experiments. Using a bacterial-fungal co-culture as a model system, this approach enabled rapid annotation of coproporphyrin III directly from culture. An untargeted precursor selection workflow was also developed within SCiLS Lab to streamline iprm-PASEF method generation and pseudo-untargeted imaging analyses.Finally, Chapter 4 demonstrates the application of MALDI-TIMS-MS as a high-throughput platform for screening engineered biocatalysts that generate isomeric products. By leveraging ion mobility separation, a workflow was developed to resolve and compare kainic acid and kainic acid lactone abundance directly from genetically diversified bacterial colonies. Screening of 1,054 genetically diversified KabC variants identified seven enzymes with improved kainic acid lactone production while maintaining favorable expression and enhanced stability, providing new tools for investigating structure-function relationships and advancing biocatalyst engineering.Collectively, this dissertation demonstrates how multidimensional, high-throughput mass spectrometry can accelerate discovery across multiple stages of the natural products pipeline. By integrating rapid molecular annotation, spatial metabolomics, and enzyme screening, these approaches establish versatile analytical platforms that expand the speed, scale, and depth of biochemical investigation.