THE SWEET PROTEIN BRAZZEIN AND ITS INTERACTION WITH THE HUMAN TASTE RECEPTOR

甜蛋白火及其与人类味觉受体的相互作用

• 批准号: 8168982
• 负责人: FARIBA M ASSADI-PORTER
• 金额: $ 2.9万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168982
• 关键词: Address; Affect; Binding; C-terminal; Carbohydrates; Cellular Assay; Computer Retrieval of Information on Scientific Projects Database; Cysteine-Rich Domain; Diabetes Mellitus; Disease; Funding; Grant; Heating; Human; In Vitro; Institution; Ligand Binding; Modeling; Molecular; Monitor; Mutagenesis; Mutation; NMR Spectroscopy; Obesity; Primates; Property; Proteins; Public Health; Research; Research Personnel; Resources; Signal Transduction; Source; Surface; Sweetening Agents; Taste Perception; United States National Institutes of Health; Venus Flytrap; extracellular; interest; mutant; receptor; receptor binding; receptor sensitivity; small molecule; sweet receptor; sweet taste perception

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Obesity as a major public health issue has increased the interest in low-calorie, natural sweeteners. A prime candidate is the 53-residue, heat stable, protein, brazzein, which contains no carbohydrate and is perceived as sweet tasting only by old world primates and humans. Through mutagenesis we have determined that three major regions near N- and C-terminal domains and Loop43 region are critical for heterodimeric sweet receptor binding and/or activity; mutations in other regions appear to affect sweetness by indirect conformational changes, as detected by NMR spectroscopy. Complementary mutagenesis studies of the sweet receptor indicate that brazzein interacts with specific residues in the Cysteine-rich domain (CRD) of the T1R3 subunit. In addition, modeling and receptor mutagenesis studies have identified a major binding surface for brazzein on the "VFTM" (Venus flytrap ligand binding extracellular module) of T1R2 subunit. This large interaction surface on the receptor distiguishes the mode of action of brazzein from those of small molecule sweeteners. Specific contributions of T1R2 resdidues in determining the differential sensitivity of the receptor to brazzein remain to be discovered. We propose three specific aims: 1. In vitro cellular assays of sweet protein-receptor interactions. 2. To use NMR spectroscopy to investigate the conformational and dynamic requirements in brazzein for its interaction and activation of the sweet taste receptor. These studies will serve to identify changes that correlate with fuctional properties. 3. To monitor binding of brazzein and its mutants to T1R2/T1R3 sweet receptor and its mutants by STDD-NMR spectroscopy. These results will contribute to accurately define the essential molecular features responsible for the brazzein-sweet receptor interaction and the resulting signal transduction non-caloric sweeteners as an approach to help address diabetes and related disorders.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 肥胖症作为一个主要的公共卫生问题，增加了对低热量，天然甜味剂的兴趣。主要的候选人是53个残留，热稳定，蛋白质，brazzein，其中不含碳水化合物，只有旧世界的灵长类动物和人类才被视为甜美的品尝。通过诱变，我们确定了N和C末端结构域和Loop43区域附近的三个主要区域对于异二聚体甜受体结合和/或活性至关重要。如NMR光谱检测，其他区域的突变似乎通过间接构象变化影响甜度。甜蜜受体的互补诱变研究表明，Brazzein与T1R3亚基的富含半胱氨酸域（CRD）中的特定残基相互作用。此外，建模和受体诱变研究已经确定了T1R2亚基的“ VFTM”（Venus flytrap配体结合细胞外模块）上Brazzein的主要结合表面。受体上的这种大相互作用表面使Brazzein的作用方式从小分子甜味剂的作用模式中汲取了动作。 T1R2重塑在确定受体对Brazzein的差异敏感性方面的特定贡献仍有待发现。我们提出了三个特定目的：1。甜蛋白质受体相互作用的体外细胞测定。 2。使用NMR光谱研究Brazzein中的构象和动态需求，以与甜味受体的相互作用和激活。这些研究将有助于确定与抗性特性相关的变化。 3。通过STDD-NMR光谱法监测Brazzein及其突变体与T1R2/T1R3甜受体及其突变体的结合。这些结果将有助于准确定义负责Brazzein-Sweet受体相互作用的基本分子特征，以及所得的信号转导非热甜味剂作为帮助解决糖尿病和相关疾病的方法。