Interdependent domains controlling the enzymatic activity of mitogen-activated protein kinase kinase 1
Interdependent domains controlling the enzymatic activity of mitogen-activated protein kinase kinase 1
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DOI：
10.1021/bi961854s
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发表时间：
1996-12-03
期刊：
影响因子：
2.9
通讯作者：
Ahn, NG
中科院分区：
文献类型：
--
作者：
Mansour, SJ;Candia, JM;Ahn, NG
人促分裂原活化蛋白激酶激酶1（MKK 1）的激活是通过调节环内Ser 218和Ser 222的磷酸化实现的。通过用天冬氨酸/谷氨酸替换这些残基来实现部分活化。通过在调节环中引入四个酸性残基取代获得更高的活性，表明环中的酸性残基通过引入负电荷稳定活性构型。MKK 1的活化也通过删除共有催化核心N-末端的残基44-51来实现。虽然在该片段内的残基被丙氨酸取代不影响活性，但脯氨酸残基的引入提高了激酶活性，表明活化是由残基44-51内的二级结构扰动引起的。假底物抑制，一种常见的激酶调节机制，在此过程中不起作用。酸性取代和N-末端缺失均增加V-max、V/K-m、K-ERK 2和V/K-m、K-ATP，这也在野生型MKK 1磷酸化后观察到。这些稳态速率参数的协同增强发生在组合突变后，表明诱变诱导的构象变化一起模仿磷酸化后看到的那些。
The activation of human mitogen-activated protein kinase kinase 1 (MKK1) is achieved by phosphorylation at Ser218 and Ser222 within a regulatory loop. Partial activation was achieved by replacing these residues with aspartic/glutamic acid. Higher activity was obtained by introducing four acidic residue substitutions in the regulatory loop, indicating that acidic residues in the loop stabilize an active configuration by the introduction of negative charge. Activation of MKK1 is also achieved by deleting residues 44-51, N-terminal to the consensus catalytic core. Although substitution of residues within this segment by alanine does not affect activity, introduction of proline residues elevates kinase activity, indicating that activation results from perturbation of secondary structure within residues 44-51. Pseudosubstrate inhibition, a commonly observed mechanism of kinase regulation, is not operative in this process. Both the acidic substitutions and the N-terminal deletion increase V-max, V/K-m,K-ERK2, and V/K-m,K-ATP, as is also observed following phosphorylation of wild-type MKK1. A synergistic enhancement of these steady-state rate parameters occurs upon combining the mutations, suggesting that conformational changes induced by mutagenesis together mimic those seen upon phosphorylation.