McKay D M, Shaw C, Thim L, Johnston C F, Halton D W, Fairweather I, Buchanan K D
Department of Medicine, Queen's University of Belfast, Northern Ireland.
Regul Pept. 1990 Dec 10;31(3):187-97. doi: 10.1016/0167-0115(90)90005-h.
Using an antiserum directed against the highly-conserved C-terminal hexapeptide amide of mammalian pancreatic polypeptide (PP), numerous immunoreactive endocrine cells were identified within the pancreas of the European common frog, R. temporaria. An acidified ethanolic extract of pancreatic tissue (0.859 g, n = 35) contained 26.2 nmol equivalents/g of tissue. Gel permeation chromatography of the extract resolved a single peak of immunoreactivity co-eluting with synthetic bovine PP standard. Reverse phase HPLC of this material resolved a single peak of immunoreactivity which was purified to homogeneity following chromatography on a semipreparative C-18 column and an analytical C-8 column. Plasma desorption mass spectrometry (PDMS) of the purified peptide resolved a single component with a molecular mass of 4240.9 Da. Direct gas phase sequencing established the sequence of the first 26 residues. Following incubation of the peptide with endopeptidase Asp-N and direct application of the digest to the sequencer, the entire primary structure of the peptide was established as: APSEPHHPGDQATQDQLAQYYSDLYQYITFVTRPRF. The derived molecular mass of this peptide, incorporating a C-terminal amide, was 4240.6 Da which is entirely consistent with that obtained by PDMS.
使用针对哺乳动物胰多肽（PP）高度保守的C末端六肽酰胺的抗血清，在欧洲普通青蛙（R. temporaria）的胰腺中鉴定出许多免疫反应性内分泌细胞。胰腺组织的酸化乙醇提取物（0.859 g，n = 35）含有26.2 nmol当量/克组织。提取物的凝胶渗透色谱法分离出一个与合成牛PP标准品共洗脱的免疫反应性单峰。该物质的反相HPLC分离出一个免疫反应性单峰，在半制备C-18柱和分析C-8柱上色谱后纯化至同质。纯化肽的等离子体解吸质谱（PDMS）解析出一个分子量为4240.9 Da的单一成分。直接气相测序确定了前26个残基的序列。将该肽与天冬氨酸内肽酶-N孵育并将消化物直接应用于测序仪后，该肽的整个一级结构确定为：APSEPHHPGDQATQDQLAQYYSDLYQYITFVTRPRF。该肽的推导分子量，包含C末端酰胺，为4240.6 Da，与通过PDMS获得的结果完全一致。
