CONSTITUTIVE ACTIVATION OF MITOGEN-ACTIVATED PROTEIN KINASE-ACTIVATED PROTEIN-KINASE-2 BY MUTATION OF PHOSPHORYLATION SITES AND AN A-HELIX MOTIF
CONSTITUTIVE ACTIVATION OF MITOGEN-ACTIVATED PROTEIN KINASE-ACTIVATED PROTEIN-KINASE-2 BY MUTATION OF PHOSPHORYLATION SITES AND AN A-HELIX MOTIF
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DOI：
10.1074/jbc.270.45.27213
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发表时间：
1995-11-10
影响因子：
4.8
通讯作者：
GAESTEL, M
中科院分区：
文献类型：
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作者：
ENGEL, K;SCHULTZ, H;GAESTEL, M
最近描述的丝裂原活化蛋白激酶（MAPK）的下游靶标是MAPK活化蛋白（MAPKAP）激酶2，其已被证明负责小热休克蛋白磷酸化。我们使用重组MAPKAP激酶2融合蛋白分析了MAPK磷酸化激活MAPKAP激酶2的机制，p44（MAPK）和p38/40（MAPK）在体外和使用表位标记的MAPKAP激酶2在热休克的NIH 3 T3细胞中。它表明，除了已知的磷酸化的苏氨酸残基的羧基末端的催化结构域，Thr-317，MAPKAP激酶2在体外和体内的激活是依赖于磷酸化的第二个苏氨酸残基，Thr-205，这是位于催化结构域内，这是高度保守的几种蛋白激酶。MAPKAP激酶2的组成型活化是通过用谷氨酸取代这两个苏氨酸残基获得的。MAPKAP激酶2的组成型活化形式也是通过缺失含有Thr-317和A-螺旋基序的羧基末端区域或通过取代A-螺旋的保守残基获得的，这些数据表明MAPKAP激酶2激活的双重机制，通过催化结构域内的Thr-205的磷酸化和催化结构域外的Thr-317的磷酸化，涉及自抑制A-螺旋基序。
A recently described downstream target of mitogen-activated protein kinases (MAPKs) is the MAPK-activated protein (MAPKAP) kinase 2 which has been shown to be responsible for small heat shock protein phosphorylation, We have analyzed the mechanism of MAPKAP kinase 2 activation by MAPK phosphorylation using a recombinant MAPKAP kinase 2-fusion protein, p44(MAPK) and p38/40(MAPK) in vitro and using an epitope-tagged MAPKAP kinase 2 in heat-shocked NIH 3T3 cells. It is demonstrated that, in addition to the known phosphorylation of the threonine residue carboxyl-terminal to the catalytic domain, Thr-317, activation of MAPKAP kinase 2 in vitro and in vivo is dependent on phosphorylation of a second threonine residue, Thr-205, which is located within the catalytic domain and which is highly conserved in several protein kinases. Constitutive activation of MAPKAP kinase 2 is obtained by replacement of both of these threonine residues by glutamic acid, A constitutively active form of MAPKAP kinase 2 is also obtained by deletion of a carboxyl-terminal region containing Thr-317 and the A-helix motif or by replacing the conserved residues of the A-helix, These data suggest a dual mechanism of MAPKAP kinase 2 activation by phosphorylation of Thr-205 inside the catalytic domain and by phosphorylation of Thr-317 outside the catalytic domain involving an autoinhibitory A-helix motif.