Adebanjo O A, Koval A, Moonga B S, Wu X B, Yao S, Bevis P J, Kumegawa M, Zaidi M, Sun L
Mount Sinai Bone Program, Mount Sinai School of Medicine, New York, New York 10029, USA.
Biochem Biophys Res Commun. 2000 Jul 14;273(3):884-9. doi: 10.1006/bbrc.2000.3041.
We report the molecular cloning and functional characterization of a novel member of the CD38 family of cyclic ADP-ribose (cADPr)-generating cyclases. We cloned a cDNA insert that encoded a 298-amino-acid-long protein (M(w) approximately 39 kDa). The predicted protein displayed 69, 61, and 58% similarity, respectively, to mouse, rat, and human CD38. Rabbit CD38 was also 28% homologous to Aplysia ADP-ribosyl cyclase and leukocyte CD157 (another ADP-ribosyl cyclase); the three cyclases shared 10 cysteine and 2 adjacent proline residues. We then transfected CD38-negative NIH3T3 cells with cDNA encoding a CD38-EGFP fusion protein. Epifluorescence microscopy showed intense EGFP fluorescence confirming CD38 expression. We finally confirmed the ADP-ribosyl cyclase activity of the expressed CD38 by measuring its ability to catalyze the cyclization of the nicotinamide adenine dinucleotide (NAD(+)) surrogate, NGD(+), to its fluorescent nonhydrolyzable derivative, cGDPr.
我们报道了一种新型环磷酸腺苷核糖（cADPr）生成环化酶CD38家族成员的分子克隆及功能特性。我们克隆了一个编码298个氨基酸长蛋白质（分子量约39 kDa）的cDNA插入片段。预测的蛋白质与小鼠、大鼠和人类CD38的相似性分别为69%、61%和58%。兔CD38与海兔ADP - 核糖基环化酶和白细胞CD157（另一种ADP - 核糖基环化酶）也有28%的同源性；这三种环化酶共有10个半胱氨酸和2个相邻的脯氨酸残基。然后我们用编码CD38 - EGFP融合蛋白的cDNA转染CD38阴性的NIH3T3细胞。落射荧光显微镜显示强烈的EGFP荧光，证实了CD38的表达。我们最终通过测量其催化烟酰胺腺嘌呤二核苷酸（NAD(+)）替代物NGD(+)环化生成其荧光不可水解衍生物cGDPr的能力，确认了所表达CD38的ADP - 核糖基环化酶活性。
