MASS SPECTROMETRY OF OPIOID NEUROPEPTIDERGIC SYSTEMS

用基质辅助激光解吸飞行时间质谱仪(MALDI-TOF MS)表征大鼠的阿片受体(OP1)。MALDI-TOF MS提供了分析完整受体所需的高质量能力，检测受体所需的高检测灵敏度，以及分析由胰酶消化产生的、将用于肽图谱和OP表征的未分解多肽混合物的能力(将使用其他酶和溴化氰)。研究完整的、完全翻译后修饰的受体是很重要的。与其他用于研究受体的方法相比，这种基于MS的方法具有高灵敏度、快速分析和最佳特异性。研究OP所需的MALDI-TOF MS的几个操作特征将被优化，包括：样品分析的速度(MALDI-TOF可以在一个平板上以每个样品大约一分钟的速度分析100个样品；该速率对于肽图谱和OP的表征是重要的)；检测灵敏度将提高到fmoL/amol水平；将通过使用源后衰变(PSD)来优化分子特异性，以获得胰蛋白酶多肽的氨基酸序列；采用延迟萃取法(DE)和反射式加速器(R)，可以优化OP1分子量(MW)的测量精度(0.1%水平)。

The rat delta opioid receptor (OP1) will be characterized with matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF MS). MALDI-TOF MS offers the high mass capability that is needed to analyze an intact receptor, the high detection sensitivity that is needed to detect a receptor, and the ability to analyze the mixture of unresolved peptides that is produced by trypsin digestion and that will be used for peptide mapping and for characterization of the OP (Other enzymes and cyanogen bromide will be used). It is important to study the intact, fully post- translationally modified receptor. Compared to the other methods that are used to study receptors, this MS-based method offers high sensitivity, rapid analysis, and optimum specificity. Several operating features of MALDI-TOF MS that are needed to study the OP will be optimized, including: the speed of sample analysis (MALDI-TOF can analyze 100 samples on a plate at a rate of ca. one minute per sample; that rate is important for peptide mapping and OP characterization); detection sensitivity will be improved to the fmol/amol level; molecular specificity will be optimized by using post-source decay (PSD) to obtain the amino acid sequence of tryptic peptides; and the mass accuracy of measuring the molecular weight (MW) of the OP1 will be optimized (0.1% level) by using delayed extraction (DE) and a reflectron (R).