The spleen protein-tyrosine kinase TPK-IIB is highly similar to the catalytic domain of p72(syk)
The spleen protein-tyrosine kinase TPK-IIB is highly similar to the catalytic domain of p72(syk)
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DOI：
10.1111/j.1432-1033.1996.0400h.x
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发表时间：
1996-09-01
期刊：
影响因子：
--
通讯作者：
Pinna, LA
中科院分区：
文献类型：
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作者：
Brunati, AM;James, P;Pinna, LA
TPK-IIB是一种蛋白激酶，主要存在于脾的胞质溶胶中，其特征在于对酸性肽底物的高比活性和低自磷酸化活性。一个突出的52-kDa组分用激酶纯化[Marin，O.，Donella-Deana，A.，Brunati，A. M.，Fischer，S. & Pinna，L. A.（1991）J.Biol.Chem.266，17798-17803]。在这里，我们表明，52 kDa的蛋白显示与米勒-迪克尔无脑蛋白（LIS 1）的序列同一性。该蛋白与任何已知的蛋白激酶无关，并且缺乏ATP结合基序。TPK-IIB的ATP结合和磷酸转移酶活性可以通过一个较小的38-kDa蛋白条带（p38/TPK-IIB）来完全解释，该蛋白条带可以在EDTA存在下通过Mono-Q/FPLC与52-kDa蛋白分离。p38/TPK-IIB的序列分析揭示了高水平的相似性，如果不是同一性：与p72（syk）的催化结构域，一个蛋白酪氨酸激酶参与造血细胞的活化。抗p72催化结构域（syk）的抗体，而不是抗其N-末端片段的抗体，与p38/TPK-IIB交叉反应。p72（syk）的肽底物特异性与p38/TPK-IIB的肽底物特异性几乎相同，这也支持将TPK-IIB分类为p72（syk）的近亲（可能是蛋白水解或选择性剪接形式）。然而，p38/TPK-IIB表现出比p72（syk）高6倍的比活性，并且似乎对肝素抑制的敏感性高50倍。因此，观察到大鼠脾脏颗粒部分对p72（syk）的Ca 2+依赖性降解伴随着催化活性增加和对肝素敏感性增加，这与过度活性的p38/TPK-IIB可能是响应细胞内Ca 2+增加而从p72（syk）蛋白水解产生的可能性一致。
TPK-IIB is a protein kinase that is predominant in the cytosol of spleen and is characterized by a high specific activity toward acidic peptide substrates and a low auto-phosphorylation activity. A prominent 52-kDa component purifies with the kinase [Marin, O., Donella-Deana, A., Brunati, A. M., Fischer, S. & Pinna, L. A. (1991) J. Biol. Chem. 266, 17798-17803]. Here we demonstrate that the 52-kDa protein displays sequence identity with the Miller-Dieker lissencephaly protein (LIS 1). The protein is not related to any known protein kinase and lacks an ATP-binding motif. The ATP binding and phosphotransferase activities of TPK-IIB can be fully accounted for by a minor 38-kDa protein band (p38/TPK-IIB) which can be separated from the 52-kDa protein by Mono-Q/FPLC in the presence of EDTA. Sequence analysis of p38/TPK-IIB reveals a high level of similarity, if not identity: with the catalytic domain of p72(syk), a protein-tyrosine kinase implicated in the activation of hematopoietic cells. Antibodies raised against the catalytic domain of p72(syk), but not antibodies raised against its N-terminal segment, cross-react with p38/TPK-IIB. The peptide substrate specificity of p72(syk) is almost identical to that of p38/TPK-IIB, which also supports the classification of TPK-IIB as a close relative (possibly a proteolytic or alternative spliced form) of p72(syk). p38/TPK-IIB, however, exhibits a specific activity which is sixfold higher than that of p72(syk) and appears to be 50-fold more sensitive to inhibition by heparin. Thus, the observation that Ca2+-dependent degradation of p72(syk) by particulate fraction of rat spleen is accompanied by increased catalytic activity and increased sensitivity to heparin would be consistent with the possibility that hyperactive p38/TPK-IIB might be proteolytically generated from p72(syk) in response to an Increase of intracellular Ca2+.