ISOZYMIC FORMS OF RAT-BRAIN CA-2+-ACTIVATED AND PHOSPHOLIPID-DEPENDENT PROTEIN-KINASE
ISOZYMIC FORMS OF RAT-BRAIN CA-2+-ACTIVATED AND PHOSPHOLIPID-DEPENDENT PROTEIN-KINASE
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DOI：
10.1073/pnas.83.22.8535
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发表时间：
1986-11-01
影响因子：
11.1
通讯作者：
HUANG, FL
中科院分区：
文献类型：
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作者：
HUANG, KP;NAKABAYASHI, H;HUANG, FL
用羟基磷灰石柱层析分离大鼠脑内钙激活和磷脂依赖蛋白激酶(EC2.7.1.37)的三种形式。这些酶被命名为I、II和III型蛋白激酶C，分子量均为82,000，在钙离子、磷脂酰丝氨酸和二油酸甘油酯存在的情况下进行自动磷酸化，并结合[~3H]佛波醇12，13-二丁酸酯。这些酶的自磷酸化导致每摩尔酶掺入1-1.5摩尔的32P。二维肽图谱分析表明，这些酶具有不同的自磷酸化位点。磷酸氨基酸分析表明，I型和III型蛋白激酶C主要在丝氨酸残基上自磷酸化，而II型蛋白激酶C型在丝氨酸和苏氨酸残基上都有自磷酸化。此外，针对三种类型的激酶混合物产生的多克隆抗体优先抑制I型和II型酶。抗I型和II型激酶的单抗只能识别它们各自的酶，而不能识别III型酶。这些结果表明，大鼠脑内存在同工酶形式的蛋白激酶C。
Three forms of rat brain Ca2+-activated and phospholipid-dependent protein kinase (EC 2.7.1.37) were separated by hydroxylapatite column chromatography. These enzymes, designated type I, II, and III protein kinase C, all have a molecular weight of 82,000, undergo autophosphorylation in the presence of Ca2+, phosphatidylserine, and diolein, and bind [3H]phorbol 12,13-dibutyrate. Autophosphorylation of these kinases resulted in an incorporation of 1-1.5mol of 32P per mol of enzyme. Two-dimensional peptide mapping analysis revealed that these kinases had different sites of autophosphorylation. Phosphoamino acid analysis showed that type I and type III protein kinase C primarily autophosphorylated at a serine residue, whereas type II kinase autophosphorylated at both serine and threonine residues. In addition, polyclonal antibodies raised against a mixture of three types of the kinase preferentially inhibited type I and type II enzymes. Monoclonal antibodies against type I and type II kinase only recognized their respective enzymes but not the type III enzyme. These results demonstrate the presence of isozymic forms of protein kinase C in rat brain.