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Molecular Drivers of FABP-mediated Endocannabinoid Signaling for Appetite Regulation

FABP 介导的内源性大麻素信号传导食欲调节的分子驱动因素

基本信息

批准号：
10683375
负责人：
RUTH E. STARK
金额：
$ 15.7万
依托单位：
CITY COLLEGE OF NEW YORK
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-15 至 2026-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10683375
关键词：
Abbreviations Address Affinity Amides Animal Model Animals Appetite Regulation Applications Grants Articulation Autoimmune Diseases Binding Binding Proteins Binding Sites Biological Assay Biophysics Cell Nucleus Chemicals Chimera organism Complex Desire for food Diffusion Dissociation Drug Design Drug Modulation Endocannabinoids Enterocytes Enzymes Esters Exhibits Fatty Acids Fluorescence Food Intake Regulation Future Genetic Transcription Glycerol Health Heart Diseases Human Hydrophobicity In Vitro Inflammation Intestines Knock-out Knockout Mice Ligand Binding Ligand Binding Domain Ligands Lipase Liver Measures Mediating Metabolic Modeling Molecular Molecular Chaperones Molecular Conformation Molecular Structure Monitor Mus NMR Spectroscopy Non-Insulin-Dependent Diabetes Mellitus Nuclear Receptors Obesity Outcome Pain Pattern Peroxisome Proliferator-Activated Receptors Peroxisome Proliferators Phenotype Physiological Protein Binding Domain Proteins Regulation Relaxation Research Risk Role Satiation Signal Transduction Signaling Molecule Site Surface Plasmon Resonance Testing Thermodynamics Thinness Titrations Underrepresented Populations United States National Institutes of Health Vertebral column Work career design endocannabinoid signaling experimental study fatty acid amide hydrolase fatty acid-binding proteins heart disease risk in vitro testing member neuroprotection preference pressure programs protein complex protein protein interaction receptor binding recruit thyroid hormone receptor associated protein 220

项目摘要

Stark, Ruth E. Molecular Drivers of FABP-mediated Endocannabinoid Signaling for Appetite Regulation Metabolic signaling by endogenous cannabinoids (ECs) is essential to the regulation of human appetite, pain, and neuroprotection. Fatty acid-binding proteins (FABPs) can either sequester the hydrophobic ECs or transport them to hydrolytic enzymes; ECs routed to the nucleus also activate the peroxisome proliferator-activated receptors (PPARs). EC levels have been correlated with obesity in knockout mice for liver (L) FABP that is co-expressed with intestinal (I) FABP in enterocytes and also for PPARa knockouts, underscoring the regulatory roles of these proteins. We will probe poorly understood EC-FABP, EC-PPAR, and FABP-EC-PPAR complexes in vitro at near-physiological concentrations, determining affinities, metabolic fates, molecular binding interfaces, and conformational changes to test mechanistic hypotheses regarding EC signaling. Trainees at multiple career stages, including those recruited from underrepresented groups in STEM, will be integrally involved in this research program. Specific questions to be addressed are as follows: (1) How do ECs choose between FABP chaperones to produce obese or lean outcomes? The possibility that ECs are delivered by LFABP to hydrolytic enzymes for metabolic breakdown rather than sequestered by IFABP in the enterocyte will be tested enzymatically, whereas the rationale for LFABP’s diminished affinity will be explored using high- pressure solution-state NMR to identify energetically favored candidate sites for binding. (2) Could transcriptional activity be driven by EC binding preferences for LFABP vs. PPARa? The PPARa ligand-binding domain (PPARa_LBD) will be purified, rigorously delipidated, and tested in vitro for EC- modulated transcriptional activity. EC binding affinities will be compared for PPARa_LBD and FABPs. (3) Could transcriptional activity be driven by EC-modulated FABP-PPAR collisions that cause conformational changes? To determine the interactions involved in the proposed FABP-mediated EC activation of PPARa transcriptional function, we will first use surface plasmon resonance to measure the binding affinity of LFABP-PPARa_LBD protein complexes, on their own and in the presence of ligands with a range of known activation efficacies. The site-specific collision-associated impact on the LFABP partner will be probed by solution-state NMR spectroscopy of the [U-15N]-enriched protein, using chemical shift perturbations of each backbone NH resonance to define the binding interface with PPARa_LBD, any allosteric structural changes that occur upon complex formation, and their modulation by EC ligands. Taken together, these experiments will advance our understanding of (macro)molecular networks that function to achieve metabolic signaling by ECs involved in appetite and pain regulation, thereby advancing our understanding of health risks related to obesity and inflammation. This understanding can guide the design of drugs that modulate these biomedically important ligand-protein and protein-protein interactions. NIH SuRE Grant Proposal

fabp介导的内源性大麻素信号在食欲调节中的分子驱动