Kim Tack-Joong, Mitsutake Susumu, Kato Mariko, Igarashi Yasuyuki
Department of Biomembrane and Biofunctional Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Kita 12, Nishi 6, Kita-ku, Sapporo 060-0812, Japan.
FEBS Lett. 2005 Aug 15;579(20):4383-8. doi: 10.1016/j.febslet.2005.06.079.
Ceramide kinase (CERK) converts ceramide (Cer) to ceramide-1-phosphate (C1P), a newly recognized bioactive molecule capable of regulating diverse cellular functions. The N-terminus of the CERK protein encompasses a sequence motif known as a pleckstrin homology (PH) domain. However, little is known regarding the functional roles of this domain in CERK. In this study, we have demonstrated that the PH domain of CERK is essential for its enzyme activity. Using site-directed mutagenesis, we have further determined that Leu10 in the PH domain has an important role in CERK activity. Replacing this residue with a neutral alanine or isoleucine, caused a dramatic decrease in CERK activity to 1% and 29%, respectively, compared to CERK, but had no effect on substrate affinity. The study presented here suggests that the PH domain of CERK is not only indispensable for its activity but also act as a regulator of CERK activity.
神经酰胺激酶（CERK）将神经酰胺（Cer）转化为神经酰胺-1-磷酸（C1P），C1P是一种新发现的能够调节多种细胞功能的生物活性分子。CERK蛋白的N端包含一个称为普列克底物蛋白同源（PH）结构域的序列基序。然而，关于该结构域在CERK中的功能作用知之甚少。在本研究中，我们证明了CERK的PH结构域对其酶活性至关重要。通过定点诱变，我们进一步确定PH结构域中的Leu10在CERK活性中起重要作用。将该残基替换为中性丙氨酸或异亮氨酸，与CERK相比，CERK活性分别急剧下降至1%和29%，但对底物亲和力没有影响。此处提出的研究表明，CERK的PH结构域不仅对其活性不可或缺，而且还作为CERK活性的调节剂。
