Supramolecular complexes ranging from processing bodies to focal adhesion sites are increasingly found to be common elements of cellular structure and function. The purinosome is a recently discovered supramolecular protein complex that regulates, both temporally and spatially, the metabolism of purine nucleotides. Because of the fundamental significance of purine biosynthesis, the novelty of this type of spatiotemporal regulation, and the importance of this pathway as a drug target, there is a critical need to elucidate the mechanisms that dictate purinosome structure and function. Little is known about how such structures are formed, regulated or trafficked, nor is there a clear understanding of how these systems control metabolic flux. The long term goal of my research program is to understand how transient, supramolecular protein complexes regulate cellular metabolism. The overall objectives for the proposed funding period are to 1) define the structural features and extrinsic factors that control purinosome function, 2) quantify the kinetic and metabolic advantages that this protein structure provides, and 3) identify functional associations that purinosomes make with other cellular structures. The central hypothesis underlying these studies is that purinosome proteins undergo structural changes, in response to external signals, which drive the assembly process. In addition, we hypothesize that purinosomes are actively trafficked along microtubules, in response to specific signals, towards certain cellular structures such as the nucleus or plasma membrane. This level of control enables cells to specifically upregulate the production of purines and metabolic intermediates at a specific cellular locus. The rationale for the proposed research is that the purinosome is an important regulatory mechanism for the biosynthesis of purines, the purine biosynthetic pathway is critical to life, and is a clinically validated drug target. Thus, a better understanding of the purinosome will provide a clearer picture of overall nucleotide metabolism with potential to translate into more selective and potent antimetabolite drugs. The approach that we are taking is innovative, in the applicant?s opinion, because it departs from the status quo by integrating a suite of interdisciplinary tools to probe the system at multiple time and length scales. This will enable us to directly link molecular determinants of function with the corresponding biological outputs at physiologically relevant time and length scales. The outcome of these studies will be the elucidation of key, physiologically relevant and potentially druggable, interactions central to purinosome function, a mechanistic model that is likely generalizable to similar protein structures, and a quantitative determination of in vitro enzymatic and in vivo metabolic effects of this structure. The proposed research is significant, because it will provide sorely needed details of the structure, function and mechanism of a new paradigm in metabolic regulation ? the dynamic and controllable assembly of a macromolecular protein complex of metabolic enzymes. Ultimately, the elucidation of the mechanistic underpinnings of purinosome function will help guide the development of improved anti-cancer, anti-viral and anti-inflammatory treatments.
The proposed research is relevant to human health because purine metabolism is a fundamental process necessary for all life, and the de novo biosynthesis of purines is a validated drug target. The proposed studies to elucidate the molecular details and mechanistic underpinnings of the purinosome are, therefore, relevant to the mission of the NIH and NIGMS, in that they will advance our understanding of biological processes and lay the foundation for advances in the treatment of human disease.
| Shek, Roger; Dattmore, Devon A; Stives, Devin P et al. (2017) Structural and Functional Basis for Targeting Campylobacter jejuni Agmatine Deiminase To Overcome Antibiotic Resistance. Biochemistry 56:6734-6742 |
