Lacapère Jean-Jacques, Boulla Geneviève, Lund Frances E, Primack Julie, Oppenheimer Norman, Schuber Francis, Deterre Philippe
INSERM U410, Faculté de Médecine Xavier Bichat, 16 rue Henri Huchard, 75018 Paris, France.
Biochim Biophys Acta. 2003 Nov 3;1652(1):17-26. doi: 10.1016/j.bbapap.2003.07.002.
The lymphoid surface antigen CD38 is a NAD(+)-glycohydrolase that also catalyzes the transformation of NAD(+) into cyclic ADP-ribose, a calcium mobilizing second messenger. In addition, ligation of CD38 by antibodies triggers signaling in lymphoid cells. Since the cytoplasmic tail of CD38 is dispensable for this latter property, we have previously proposed that CD38-mediated receptor signal transduction might be regulated by its conformational state. We have now examined the molecular changes of this protein during its interaction with NAD(+) by measuring the intrinsic fluorescence of CD38. We have shown that addition of the substrate produced a dramatic decrease in the fluorescence of the catalytically active recombinant soluble ectodomain of murine CD38. Analysis of this event revealed that the catalytic cycle involves a state of the enzyme that is characterized by a low fluorescence which, upon substrate turnover, reverts to the initial high intrinsic fluorescence level. In contrast, non-hydrolyzable substrates trap CD38 in its altered low fluorescence state. Studies with the hydrophilic quencher potassium iodide revealed that the tryptophan residues that are mainly involved in the observed changes in fluorescence, are remote from the active site. Similar data were also obtained with human CD38, indicating that studies of intrinsic fluorescence will be useful in monitoring the transconformation of CD38 from different species. Together, these data demonstrate that CD38 undergoes a reversible conformational change after substrate binding, and suggest a mechanism by which this change could alter interactions with different cell-surface partners.
淋巴样表面抗原CD38是一种NAD(+) - 糖水解酶，它还能催化NAD(+)转化为环ADP - 核糖，一种可动员钙的第二信使。此外，抗体与CD38的结合会触发淋巴样细胞中的信号传导。由于CD38的细胞质尾巴对于后一种特性是可有可无的，我们之前曾提出CD38介导的受体信号转导可能受其构象状态的调节。我们现在通过测量CD38的内在荧光来研究该蛋白在与NAD(+)相互作用过程中的分子变化。我们已经表明，添加底物会使小鼠CD38具有催化活性的重组可溶性胞外结构域的荧光显著降低。对这一事件的分析表明，催化循环涉及一种酶的状态，其特征是荧光较低，在底物周转时，会恢复到初始的高内在荧光水平。相反，不可水解的底物会使CD38处于改变后的低荧光状态。用亲水性淬灭剂碘化钾进行的研究表明，主要参与观察到的荧光变化的色氨酸残基远离活性位点。用人CD38也获得了类似的数据，表明对内在荧光的研究将有助于监测来自不同物种的CD38的构象转变。总之，这些数据表明CD38在底物结合后会发生可逆的构象变化，并提出了一种这种变化可能改变与不同细胞表面伙伴相互作用的机制。
