Villa-Moruzzi E
Arch Biochem Biophys. 1986 May 15;247(1):155-64. doi: 10.1016/0003-9861(86)90544-8.
Phosphorylase phosphatase purified from the protein-glycogen complex of rabbit muscle has a Mr of 34,000 by gel filtration and migrates as a single band of Mr 38,000 on sodium dodecyl sulfate gel electrophoresis, i.e., of the same size as the catalytic subunit of the sarcoplasmic complex of type 1 phosphatase (L. M. Ballou, D. L. Brautigan, and E. H. Fischer (1983) Biochemistry 22, 3393-3399). The enzyme, called PG-Ea, has a specific activity of 12,000 units/mg of protein and is essentially fully active, displaying at most a 20% further increase in activity on treatment with trypsin. As in the case of the catalytic subunit of the sarcoplasmic enzyme, tryptic attack decreases the size of PG-Ea to 33,000. PG-Ea is completely inhibited by the modulator protein (inhibitor 2) after formation of a complex of Mr 70,000. On incubation of this complex at 30 degrees C, the catalytic subunit is converted (t 1/2 = 30 min) to an inactive form (Ei) that can be reactivated by the protein kinase FA (J. R. Vandenheede, S.-D. Yang, J. Goris, and W. Merlevede (1980) J. Biol. Chem. 255, 11,768-11,774) or to a lower extent by trypsin-Mn2+. Also the trypsinized PG-Ea is inhibited by inhibitor 2, it forms with this a Mr 70,000 complex and undergoes an even faster (t 1/2 = 10 min) conversion to an Ei form that can be reactivated by the kinase FA or to a lower extent by trypsin-Mn2+. Enzymological comparison of PG-Ea and trypsinized PG-Ea with the FA-activated, isolated catalytic subunit of the sarcoplasmic phosphatase (called EaFA, E. Villa-Moruzzi, L. M. Ballou, and E. H. Fischer (1984) J. Biol. Chem. 259, 5857-5863) and with its trypsinized form shows several similarities. The most relevant of these is the very specific interaction with inhibitor 2, that takes to inactivation and allows the following reactivation by FA or by trypsin-Mn2+. However, the inactivation-reactivation patterns show also some differences, namely (i) the t 1/2 of the conversion to Ei is longer for PG-Ea than for EaFA, and (ii) the reactivation of PG-Ea and trypsinized PG-Ea with trypsin-Mn2+ is only partial. Altogether the similarity but not identity of PG-Ea and EaFA would suggest that they are either two different conformations of the same molecule or two isozymes.
从兔肌蛋白质 - 糖原复合物中纯化得到的磷酸化酶磷酸酶，通过凝胶过滤测得其相对分子质量为34,000，在十二烷基硫酸钠凝胶电泳上迁移为一条相对分子质量为38,000的单带，即与1型磷酸酶肌浆复合物的催化亚基大小相同（L. M. 巴卢、D. L. 布劳蒂根和E. H. 费舍尔（1983年）《生物化学》22卷，3393 - 3399页）。这种酶称为PG - Ea，比活性为12,000单位/毫克蛋白质，基本完全有活性，用胰蛋白酶处理后活性最多再增加20%。与肌浆酶的催化亚基情况一样，胰蛋白酶作用会使PG - Ea的大小降至33,000。形成相对分子质量为70,000的复合物后，PG - Ea会被调节蛋白（抑制剂2）完全抑制。将该复合物在30℃孵育时，催化亚基会转变（半衰期 = 30分钟）为一种无活性形式（Ei），可被蛋白激酶FA重新激活（J. R. 范登海德、S. - D. 杨、J. 戈里斯和W. 默勒维德（1980年）《生物化学杂志》255卷，11,768 - 11,774页），或者在较低程度上被胰蛋白酶 - Mn²⁺重新激活。胰蛋白酶处理后的PG - Ea也会被抑制剂2抑制，与之形成相对分子质量为70,000的复合物，并更快地（半衰期 = 10分钟）转变为Ei形式，可被激酶FA重新激活，或者在较低程度上被胰蛋白酶 - Mn²⁺重新激活。将PG - Ea和胰蛋白酶处理后的PG - Ea与肌浆磷酸酶的FA激活的分离催化亚基（称为EaFA，E. 维拉 - 莫鲁齐、L. M. 巴卢和E. H. 费舍尔（1984年）《生物化学杂志》259卷，5857 - 5863页）及其胰蛋白酶处理形式进行酶学比较，发现有几个相似之处。其中最相关的是与抑制剂2的非常特异性的相互作用，这种相互作用导致失活，并允许随后被FA或胰蛋白酶 - Mn²⁺重新激活。然而，失活 - 重新激活模式也存在一些差异，即（i）PG - Ea转变为Ei的半衰期比EaFA的长，（ii）用胰蛋白酶 - Mn²⁺对PG - Ea和胰蛋白酶处理后的PG - Ea进行重新激活只是部分的。总体而言，PG - Ea和EaFA的相似但不同表明它们要么是同一分子的两种不同构象，要么是两种同工酶。
