Jiménez C R, Li K W, Dreisewerd K, Mansvelder H D, Brussaard A B, Reinhold B B, Van der Schors R C, Karas M, Hillenkamp F, Burbach J P, Costello C E, Geraerts W P
Graduate School Neurosciences Amsterdam, Research Institute Neurosciences Vrije Universiteit, Faculty of Biology, De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands.
Proc Natl Acad Sci U S A. 1997 Aug 19;94(17):9481-6. doi: 10.1073/pnas.94.17.9481.
We have established a differential peptide display method, based on a mass spectrometric technique, to detect peptides that show semiquantitative changes in the neurointermediate lobe (NIL) of individual rats subjected to salt-loading. We employed matrix-assisted laser desorption/ionization mass spectrometry, using a single-reference peptide in combination with careful scanning of the whole crystal rim of the matrix-analyte preparation, to detect in a semiquantitative manner the molecular ions present in the unfractionated NIL homogenate. Comparison of the mass spectra generated from NIL homogenates of salt-loaded and control rats revealed a selective and significant decrease in the intensities of several molecular ion species of the NIL homogenates from salt-loaded rats. These ion species, which have masses that correspond to the masses of oxytocin, vasopressin, neurophysins, and an unidentified putative peptide, were subsequently chemically characterized. We confirmed that the decreased molecular ion species are peptides derived exclusively from propressophysin and prooxyphysin (i.e., oxytocin, vasopressin, and various neurophysins). The putative peptide is carboxyl-terminal glycopeptide. The carbohydrate moiety of the latter peptide was determined by electrospray tandem MS as bisected biantennary Hex3HexNAc5Fuc. This posttranslational modification accounts for the mass difference between the predicted mass of the peptide based on cDNA studies and the measured mass of the mature peptide.
我们基于质谱技术建立了一种差异肽段展示方法，用于检测在高盐负荷的个体大鼠神经中间叶（NIL）中呈现半定量变化的肽段。我们采用基质辅助激光解吸/电离质谱，结合单参考肽段并仔细扫描基质-分析物制剂的整个晶体边缘，以半定量方式检测未分级的NIL匀浆中存在的分子离子。对高盐负荷大鼠和对照大鼠的NIL匀浆产生的质谱进行比较，发现高盐负荷大鼠的NIL匀浆中几种分子离子种类的强度有选择性且显著降低。这些离子种类的质量与催产素、加压素、神经垂体素和一种未鉴定的假定肽段的质量相对应，随后对其进行了化学表征。我们证实，分子离子种类减少的是仅源自前促垂体素和前催产素的肽段（即催产素、加压素和各种神经垂体素）。假定的肽段是羧基末端糖肽。通过电喷雾串联质谱确定后一种肽段的碳水化合物部分为二分双触角型Hex3HexNAc5Fuc。这种翻译后修饰解释了基于cDNA研究预测的肽段质量与成熟肽段测量质量之间的质量差异。
