Abe T, Swieter M, Imai T, Hollander N D, Befus A D
Department of Microbiology and Infectious Diseases, University of Calgary, Alberta, Canada.
Eur J Immunol. 1990 Sep;20(9):1941-7. doi: 10.1002/eji.1830200911.
Peritoneal mast cells (PMC) and intestinal mucosal mast cells (IMMC) were purified from rats infected with the nematode Nippostrongylus brasiliensis. Overall protein constituents of both mast cell subtypes were analyzed by two-dimensional gel electrophoresis using either nonequilibrium pH gradient electrophoresis (NEPHGE) or isoelectric focusing (IEF) in the first dimension and SDS-PAGE (10%) in the second dimension followed by silver staining. PMC had seven dominant basic proteins (PB2-8; pI 9-9.5) with estimated molecular masses of 26 to 37 kDa, as well as 80 to 90 neutral or acidic proteins, most of which had pI 6 to 7.5 and estimated molecular masses of 20 to 100 kDa. All the basic proteins were granule-associated. Three basic proteins, PB6 (29 kDa), PB7 (28 kDa) and PB8 (RMCP I, 26 kDa), bound [3H]diisopropyl fluorophosphate (DFP), suggesting that they are serine proteases. However, only PB8 was reactive with antibodies to RMCP I. Another basic component (less than 14 kDa), perhaps a degradation product of PB6, PB7 or PB8, also bound [3H]DFP. By comparison, IMMC possessed nine basic proteins (IB1-9) and, in general, they were more acidic (pI about 8.5-9) than those of PMC. Four major basic proteins (IB6-9) were all 24 kDa but were slightly different in isoelectric points. These and another 46-kDa basic component (IB2) were reactive with antibodies to RMCP II and bound [3H]DFP. There were no other DFP-binding proteins in IMMC. In spite of remarkable differences between basic granule-associated proteins in PMC and basic proteins in IMMC, spots in the neutral-acidic range were for the most part similar in the two mast cell subsets, although quantitative differences were evident for some spots. Thus, rat mast cell populations from the peritoneal cavity and intestinal mucosa exhibit marked heterogeneity in their protein constituents with basic pI, including in their granule-associated proteins with serine protease activity.
从感染巴西日圆线虫的大鼠中纯化出腹膜肥大细胞（PMC）和肠黏膜肥大细胞（IMMC）。使用非平衡pH梯度电泳（NEPHGE）或等电聚焦（IEF）作为第一向，10%的SDS-PAGE作为第二向，随后进行银染，通过二维凝胶电泳分析两种肥大细胞亚型的总蛋白成分。PMC有7种主要的碱性蛋白（PB2 - 8；pI 9 - 9.5），估计分子量为26至37 kDa，以及80至90种中性或酸性蛋白，其中大多数pI为6至7.5，估计分子量为20至100 kDa。所有碱性蛋白都与颗粒相关。三种碱性蛋白，PB6（29 kDa）、PB7（28 kDa）和PB8（RMCP I，26 kDa），结合[3H]二异丙基氟磷酸酯（DFP），表明它们是丝氨酸蛋白酶。然而，只有PB8与RMCP I抗体反应。另一种碱性成分（小于14 kDa），可能是PB6、PB7或PB8的降解产物，也结合[3H]DFP。相比之下，IMMC有9种碱性蛋白（IB1 - 9），总体而言，它们比PMC的碱性蛋白酸性更强（pI约为8.5 - 9）。四种主要的碱性蛋白（IB6 - 9）均为24 kDa，但等电点略有不同。这些蛋白以及另一种46 kDa的碱性成分（IB2）与RMCP II抗体反应并结合[3H]DFP。IMMC中没有其他DFP结合蛋白。尽管PMC中与颗粒相关的碱性蛋白和IMMC中的碱性蛋白存在显著差异，但在两个肥大细胞亚群中，中性 - 酸性范围内的斑点在很大程度上是相似的，尽管某些斑点存在明显的定量差异。因此，来自腹腔和肠黏膜的大鼠肥大细胞群体在其碱性pI的蛋白成分上表现出明显的异质性，包括在具有丝氨酸蛋白酶活性的颗粒相关蛋白方面。
