Ohira Riki H, Dipple Katrina M, Zhang Yao-Hua, McCabe Edward R B
Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA.
Biochem Biophys Res Commun. 2005 May 27;331(1):239-46. doi: 10.1016/j.bbrc.2005.03.143.
Glycerol kinase (GK) is a key enzyme in glycerol metabolism with two alternatively spliced forms-one with an 87bp insertion corresponding to exon 18 (GK+EX18), and one lacking exon 18 (GK-EX18). We report the expression of GK+/-EX18 in various tissues and cell lines, as well as their enzymatic characteristics and subcellular localization. RT-PCR revealed differential expression in tissues and cell lines. Northern blot analysis revealed that both forms of the murine ortholog, Gyk, were highly expressed in murine heart and increased during embryonic development. K(m) values for glycerol for GK+/-EX18 were not significantly different, although GK-EX18 had a higher V(max) for glycerol. GK-EX18 had a lower K(m) and V(max) for ATP than GK+EX18. Immunofluorescence experiments showed that GK+EX18 co-localized to the mitochondria and the perinuclear region while GK-EX18 had a diffuse expression pattern. These data suggest specific and divergent roles for GK+EX18 and GK-EX18 in cellular metabolism and development.
甘油激酶（GK）是甘油代谢中的关键酶，有两种可变剪接形式——一种带有对应于外显子18的87bp插入片段（GK+EX18），另一种缺少外显子18（GK-EX18）。我们报告了GK+/-EX18在各种组织和细胞系中的表达情况，以及它们的酶学特性和亚细胞定位。逆转录聚合酶链反应（RT-PCR）显示其在组织和细胞系中的表达存在差异。Northern印迹分析表明，小鼠直系同源基因Gyk的两种形式在小鼠心脏中均高表达，并在胚胎发育过程中增加。GK+/-EX18对甘油的米氏常数（K(m)）值无显著差异，尽管GK-EX18对甘油的最大反应速度（V(max)）较高。GK-EX18对三磷酸腺苷（ATP）的K(m)和V(max)均低于GK+EX18。免疫荧光实验表明，GK+EX18共定位于线粒体和核周区域，而GK-EX18呈弥漫性表达模式。这些数据表明GK+EX18和GK-EX18在细胞代谢和发育中具有特定且不同的作用。
