Purification and properties of the native form of the purple acid phosphatase from bovine spleen.
Purification and properties of the native form of the purple acid phosphatase from bovine spleen.
复制标题
DOI：
10.1021/bi00083a010
复制
发表时间：
1993-08
期刊：
影响因子：
2.9
通讯作者：
J. Orlando;T. Zirino;B. Quirk;B. Averill
中科院分区：
文献类型：
--
作者：
J. Orlando;T. Zirino;B. Quirk;B. Averill
牛脾中的紫色酸性磷酸酶(PAP)在完整的脾组织中以约36 kDa的单一多肽形式存在。先前分离的15 kDa和23 kDa或21 kDa亚基的微异质复合体似乎是由多肽链中暴露的、高度可变的环的蛋白水解性裂解而产生的。已经获得了少量的单一多肽形式，推测是酶的天然形式；这使得它的光学和EPR光谱以及基本的动力学性质得以确定。天然形式和两亚单位形式的PAP之间最显著的区别是后者的酶活性高约3倍，这是由于Vmax的简单增加。这两种形式在光谱和化学上非常相似，似乎不同的只是在155和160位之间失去了多肽的一个高度抗原性的约5个氨基酸片段，而在NH2末端序列或碳水化合物含量方面没有区别。对已发表的序列数据的分析表明，在脾PAP序列的155-161位处存在一个暴露的高抗原环，这是PAP的一个相对普遍的特征。胰蛋白酶和胰凝乳酶都能切割牛脾PAP和子宫铁蛋白，明显地在这个区域，酶活性显著增强。
The purple acid phosphatase (PAP) from bovine spleen has been shown to exist as a single ca. 36-kDa polypeptide in intact spleen tissue. The previously isolated microheterogeneous complex of 15-kDa and 23- or 21-kDa subunits appears to arise from proteolytic cleavage of an exposed, highly variable loop in the polypeptide chain. Small amounts of a single polypeptide form, presumed to be the native form of the enzyme, have been obtained; this has permitted its optical and EPR spectra and fundamental kinetic properties to be determined. The most notable difference between the native and two-subunit forms of PAP is a ca. 3-fold higher enzymatic activity for the latter, which is due to a simple increase in Vmax. The two forms are very similar spectroscopically and chemically and appear to differ only in the loss of a highly antigenic ca. five amino acid segment of the polypeptide between positions 155 and 160 but not in NH2-terminal sequence or in carbohydrate content. Analysis of published sequence data suggests that the existence of an exposed highly antigenic loop at positions corresponding to 155-161 of the spleen PAP sequence is a relatively general feature of PAP's. Trypsin and chymotrypsin cleave both bovine spleen PAP and uteroferrin, apparently in this region, with significant enhancement of enzymatic activity.