Friday, May 23, 2025
5/23/2025
Elucidation of the chemical and biological roles of five-carbon metabolism - Project Summary Our long-term goal is to decipher the multifaceted role of five-carbon (C5) metabolism in cellular physiology and disease. Five-carbon metabolism derives from the central hub of the isoprenoid pathway, which is necessary for an array of critical bioactivities, including cell membrane integrity (e.g., cholesterol), glycoprotein synthesis (e.g., the dolichols), steroid hormone signaling (e.g., androgens, estrogens, and cortisol), and mitochondrial health (e.g., coenzyme Q). Human isoprenoids derive from the mevalonic acid (MVA) pathway, whereas many other organisms utilize the methyl erythritol phosphate (MEP) pathway. The MVA and MEP pathways both converge on the same two isomeric C5 metabolites, isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP). Therefore, IPP and DMAPP are the central five-carbon precursors for all isoprenoids in all organisms. Despite their importance, there are relatively few chemical and biological tools to directly study IPP and DMAPP and little is understood about their independent biological activity, and metabolic fate beyond incorporation into longer chain isoprenoids. Furthermore, no chemical probes or genetic tools have been developed to modulate the activity of human isopentenyl pyrophosphate isomerase (IPPI), which is necessary to produce DMAPP and maintain homeostasis of the C5 precursor pool. Levels of IPP and DMAPP are directly involved in cardiovascular disease and IPPI has recently been implicated in the progression of Alzheimer’s, and the development disorders Zellweger spectrum disorder and adrenoleukodystrophy. Extensive prior research efforts have focused on developing inhibitors for enzymes in the MVA pathway, but human IPPI has not been explored. Similarly, chemical tools developed for studying prenylation via the long-chain isoprenoids farnesyl pyrophosphate (C15) and geranylgeranyl pyrophosphate (C20) are well-established, but tools and approaches for interrogating C5 prenylation are completely lacking. The collective lack of tools and approaches for studying C5 metabolism is likely caused in part by the fact that IPP and DMAPP are cell impermeant and thus cannot be exogenously delivered to cells. We recently addressed this key barrier through the development of cell-permeant analogs of IPP and DMAPP and demonstrated their utility in human cancer cell lines and the model gram positive bacterium Bacillus subtilis. Motivated by our recent progress and proof-of-principle preliminary results supporting the research methodology, we will address unmet needs by: (1) creating the next generation cell-permeant C5 analogs and developing inhibitors and cellular tools targeting modulation of IPPI activity; (2) develop a high throughput assay for the quantification of IPP and DMAPP in cellular experiments; and (3) Combine the developed chemical biology tools with advanced analytical approaches to discover and characterize novel metabolites and biological functions of C5 metabolism. Overall, this research promises to provide versatile tools for detailed investigations of C5 metabolism, expand our scientific understanding of C5 prenylation, and shed light on potential therapeutic targets in the isoprenoid pathway.
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