Low Huiyu, Chua Chun Song, Sim Tiow-Suan
Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, Block MD4A #04-02, Singapore, Singapore.
Cell Mol Life Sci. 2009 Sep;66(18):3081-90. doi: 10.1007/s00018-009-0101-8. Epub 2009 Jul 31.
A mitogen-activated protein kinase (MAPK), Pfmap2, has been identified in Plasmodium falciparum. However, its bona fide activator remains elusive as no MAPK kinase (MAPKK) homologues have been found so far. Instead, Pfnek3, a NIMA (never in mitosis, Aspergillus)-related kinase, was earlier reported to display a MAPKK-like activity due to its activating effect on Pfmap2. In this study, the regulatory mechanism of Pfnek3 was investigated. Pfnek3 was found to possess a SSEQSS motif within its activation loop that fulfills the consensus SXXXS/T phospho-activating sequence of MAPKKs. Functional analyses of the SSEQSS motif by site-directed mutagenesis revealed that phosphorylation of residues S221 and S226 is essential for mediating Pfnek3 activity. Moreover, via tandem mass-spectrometry, residue T82 was uncovered as an additional phosphorylation site involved in Pfnek3 activation. Collectively, these results provide valuable insights into the potential in vivo regulation of Pfnek3, with residues T82, S221 and S226 functioning as phospho-activating sites.
在恶性疟原虫中已鉴定出一种丝裂原活化蛋白激酶（MAPK），即Pfmap2。然而，其真正的激活剂仍然难以捉摸，因为迄今为止尚未发现MAPK激酶（MAPKK）的同源物。相反，较早前有报道称，一种与NIMA（Aspergillus中从未有过有丝分裂）相关的激酶Pfnek3，由于其对Pfmap2的激活作用而表现出类似MAPKK的活性。在本研究中，对Pfnek3的调控机制进行了研究。发现Pfnek3在其激活环内具有一个SSEQSS基序，该基序符合MAPKKs的共有SXXXS/T磷酸化激活序列。通过定点诱变对SSEQSS基序进行功能分析表明，S221和S226残基的磷酸化对于介导Pfnek3活性至关重要。此外，通过串联质谱法，发现残基T82是参与Pfnek3激活的另一个磷酸化位点。总的来说，这些结果为Pfnek3在体内的潜在调控提供了有价值的见解，其中残基T82、S221和S226作为磷酸化激活位点发挥作用。
