Analysis of NAD-cleavage Enzymes Involved in Signal Transduction System

人细胞表面抗原CD 38，其氨基酸序列与Astrasia ADP-核糖基环化酶同源，是一种具有单跨膜结构域的46-kDa II型糖蛋白。我们先前证明了CD 38的胞外结构域表现出NAD+糖水解酶（NAD酶）活性，并且HL-60细胞中由全反式视黄酸（RA）诱导的胞外形式NAD酶活性是由于CD 38（Kontani，K.et al.，J.Biol.Chem.268：16895，1993）。CD 38不仅催化NAD^+的水解，而且还催化环状ADP-核糖的形成和水解，后者是介导细胞内Ca ^2+库释放Ca^2+的新候选物。在本研究中，我们获得了以下发现。1.除了这些酶活性，CD 38具有结合透明质酸的能力。2.用抗CD 38单克隆抗体刺激RA分化的HL-60细胞，可诱导分子量为120，000、87，000和77，000的细胞蛋白快速酪氨酸磷酸化。其中一个显著的磷酸化蛋白被鉴定为c-cbl原癌基因产物p120^<c-cbl>[3]。抗CD 38单克隆抗体能显著增强分化HL-60细胞对甲酰-Met-Leu-Phe的超氧化物生成。4. Zn^（2+）与CD 38直接相互作用，使其催化性质从NAD酶转变为ADP-核糖基环化酶，这可能是由于Zn ^（2+）阻止了水分子进入酶-底物复合物的中间体。5.虽然CD 38 mRNA的表达是通过核内RA受体介导的，但CD 38基因第一内含子中的负调控元件似乎参与了RA诱导的HL-60细胞CD 38 mRNA的表达。

The human cell surface antigen CD38, which has an amino acid sequence homologous to Aplysia ADP-ribosyl cyclase, is a 46-kDa type II glycoprotein with a single-transmembrane domain. We previously demonstrated that the extracellular domain of CD38 exhibits NAD^+ glycohydrolase (NADase) activity and that the ecto-form NADase activity induced by all-trans retinoic acid (RA) in HL-60 cells is due to CD38 (Kontani, K.et al., J.Biol.Chem.268 : 16895,1993). CD38 catalyzes not only the hydrolysis of NAD^+, but also the formation and hydrolysis of cyclic ADP-ribose, which is a novel candidate that mediates Ca^<2+> release from intracellular Ca^<2+> stores. In the present study, we obtained the following findings. 1. Besides these enzyme activities, CD38 had the ability to bind hyaluronate. 2. Stimulation of RA-differentiated HL-60 cells with anti-CD38 monoclonal antibody (mAb) induced rapid tyrosine phosphorylation of cellular proteins with the molecular weight of 120,000,87,000 and 77,000. One of the prominent phosphorylated proteins was identified as the c-cbl proto-oncogene product, p120^<c-cbl>.3. Superoxide formation in response to formyl-Met-Leu-Phe was markedly enhanced by the anti-CD38 mAb in the differentiated HL-60 cells. 4. Zn^<2+> directly interacted with CD38 to convert its catalytic properties from NADase to ADP-ribosyl cyclase, probably due to prevention of the access of water molecule to an intermediate of the enzyme-substrate complex. 5. Although the expression of CD38 mRNA was mediated through nuclear RA receptors, a negative regulatory element present in the first intron of CD38 gene appeared to be involved in the RA-induced expression of CD38 mRNA in HL-60 cells.