小鼠鞘氨醇激酶1a的C4结构域中的天冬氨酸177对鞘氨醇的识别很重要。
Asp177 in C4 domain of mouse sphingosine kinase 1a is important for the sphingosine recognition.
作者信息
Yokota Shinji, Taniguchi Yuki, Kihara Akio, Mitsutake Susumu, Igarashi Yasuyuki
机构信息
Department of Biomembrane and Biofunctional Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12, Nishi 6, Kita-ku, Sapporo 060-0812, Japan.
出版信息
FEBS Lett. 2004 Dec 3;578(1-2):106-10. doi: 10.1016/j.febslet.2004.10.081.
Sphingosine kinase (SK) is the enzyme that catalyzes the formation of sphingosine 1-phosphate (S1P). Although diverse biological functions have been reported for SK, its recognition site for its substrate sphingosine (Sph) is still unclear. We constructed various mutants of mouse sphingosine kinase 1a (mSK1a), carrying mutations in the C4 domain, which we had expected to encompass the Sph-binding site. We analyzed the influence of these mutations on the SK activity and substrate kinetics. One mutation, Asp177-->Asn177, caused a dramatic decrease in SK activity (to approximately 6% of wild type) and an increase in the Km value for Sph (10.1-->108 microM), with no change in the affinity for ATP. This result suggests that the C4 domain, especially the Asp177, is involved in the specific recognition of Sph. In this report, we are able, for the first time, to provide an account of the Sph-binding site of SK.
鞘氨醇激酶（SK）是一种催化生成1-磷酸鞘氨醇（S1P）的酶。尽管已有多种SK的生物学功能被报道，但其底物鞘氨醇（Sph）的识别位点仍不清楚。我们构建了小鼠鞘氨醇激酶1a（mSK1a）的各种突变体，这些突变体在C4结构域携带突变，我们预期该结构域包含Sph结合位点。我们分析了这些突变对SK活性和底物动力学的影响。一种突变，Asp177→Asn177，导致SK活性急剧下降（降至野生型的约6%），Sph的Km值增加（从10.1→108μM），而对ATP的亲和力没有变化。该结果表明C4结构域，尤其是Asp177，参与了对Sph的特异性识别。在本报告中，我们首次能够阐述SK的Sph结合位点。
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