Chemical Biology Approaches to Studying Collagen IV Stability

研究胶原蛋白 IV 稳定性的化学生物学方法

• 批准号: 10723042
• 负责人: Jason E Gestwicki
• 金额: $ 43.57万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10723042
• 关键词: Affinity; Anabolism; Animals; Automobile Driving; Basement membrane; Binding; Biochemical; Biogenesis; Biological Assay; Biology; Blindness; C-terminal; Cartilage; Cataract; Cell Line; Cell secretion; Cells; Characteristics; Chemicals; Collagen; Collagen Gene; Collagen Type IV; Corneal dystrophy; Defect; Diagnosis; Disease; Drug Combinations; Ehlers-Danlos Syndrome; Endoplasmic Reticulum; Extracellular Matrix; Extracellular Space; Family; Future; Genetic; Genetic Diseases; Glaucoma; Glues; Goals; Gould Syndrome; Hearing; Hereditary nephritis; Individual; Kidney; Laboratories; Length; Methods; Microphthalmos; Missense Mutation; Modeling; Molecular; Muscle Weakness; Mutagenesis; Mutation; N-terminal; Nonsense Mutation; Organ; Osteogenesis Imperfecta; Pathway interactions; Penetrance; Persons; Pharmaceutical Preparations; Phenotype; Physiologic Intraocular Pressure; Play; Post-Translational Protein Processing; Process; Protein Engineering; Proteins; Protomer; Recombinants; Role; Signaling Molecule; Site; Skin; Symptoms; System; Technology; Testing; Tissues; Variant; Work; autosome; biochemical model; bone; cell motility; design; drug discovery; drug-like compound; experience; eye dryness; gene therapy; high throughput screening; improved; innovation; mechanical properties; mouse model; preclinical study; rare genetic disorder; self assembly; skeletal; small molecule; tool; triple helix

PROJECT SUMMARY/ABSTRACT Collagens are essential components of the extra-cellular matrix and basement membranes, where they serve to maintain tissue integrity, facilitate cell migration, and organize signaling molecules. To achieve these activities, mature collagens are secreted from cells as stable trimers. Each collagen protomer is composed of three domains: a C-terminal, non-collagenous (NC1) domain, a collagen domain and an N-terminal domain (NTD). Newly synthesized collagen proteins undergo a unique “C-to-N” folding pathway in the endoplasmic reticulum (ER), in which individual, C-terminal NC1 domains first assemble into trimers prior to collagen triple helix formation. Collagens are also heavily modified by post-translational modifications (PTMs) prior to secretion of mature collagen trimers to the extracellular space. Collagen-associated disorders (CADs), such as Osteogenesis Imperfecta, Ehlers-Danlos syndrome (EDS) and Alport’s syndrome, are relatively common (1:5,000 to 1:10,000) genetic diseases that most commonly arise from mutations in collagen genes. Because of the key roles played by collagens in ocular tissues, vision loss is a common feature of CADs. For example, ocular dysgenesis is a feature of Gould Syndrome, a rare genetic disorder associated with mutations in type IV collagen a1 (COL4A1) and type IV collagen a2 (COL4A2). We propose that stabilizing NC1 interactions with genetic (e.g., gene therapy) or chemical (e.g., molecular glues) approaches is a potential, new way to treat Gould Syndrome. Specifically, we envision that stabilization of COL4A1-COL4A2 contacts will partially overcome the impact of missense or nonsense mutations by driving assembly of type IV collagen [a1a2a2(IV)] heterotrimers. To explore this idea, we used computational protein design to create a “stabilizer” variant, COL4A2S150W, in which residue packing at the COL4A2-COL4A1 interface is optimized. Indeed, we found that expression of COL4A2S150W enhanced biogenesis of collagen [a1a2a2(IV)] by 7-fold in a quantitative, cell-based model. Now, we are poised to (Specific Aim 1) screen Gould Syndrome- associated mutations to reveal which ones can be partially corrected in cell-based and biochemical models. Then, we will (Specific Aim 2) conduct high throughput chemical screens, which leverage our innovative, cell- based assays and “stabilizer” COL4A2S150W tool, to identify drug-like molecules that promote collagen [a1a2a2(IV)] biogenesis. Together, we expect these studies to reveal the relationship between NC1 affinity and collagen secretion in models of Gould Syndrome. We also hope to produce starting points for new Gould Syndrome treatments and provide a framework for potentially treating other CADs.

项目总结/文摘