Abstract：
The quantitative and qualitative biochemical description of molecular interactions is fundamental to the study of ligand/receptor pairs and their structure/function relationships. Bioactive peptides often are active at (sub-)nanomolar concentrations, indicating they have a high affinity for their sites of action, notably binding sites on receptors. Since such receptor proteins are commonly of low abundance, highly sensitive detection methods are required to study these ligand/receptor interactions. We present a protocol for an inexpensive luminescence-based detection setup in which the peptide ligand of interest is extended with the 11-amino acid HiBiT tag. This tag can be quantified easily down to fmol amounts by its ability to reconstitute the enzymatic activity of LgBiT, a truncated version of the Oplophorus gracilirostris luciferase.
摘要：
分子相互作用的定量和定性生化描述是研究配体/受体对及其结构/功能关系的基础。生物活性肽通常在（亚）纳摩尔浓度下具有活性，表明他们对行动地点有很高的亲和力，特别是受体上的结合位点。由于这种受体蛋白通常丰度较低，需要高灵敏度的检测方法来研究这些配体/受体相互作用。我们提出了一种廉价的基于发光的检测装置的方案，其中感兴趣的肽配体用11个氨基酸的HiBiT标签延伸。该标签可以通过其重建LgBiT酶活性的能力轻松定量至fmol量，章鱼荧光素酶的截短版本。
