Mulero J J, Yeung G, Nelken S T, Ford J E
Functional Genomics Department, Hyseq Inc., Sunnyvale, California 94086, USA.
J Biol Chem. 1999 Jul 16;274(29):20064-7. doi: 10.1074/jbc.274.29.20064.
The human ecto-apyrase gene family consists of five reported members (CD39, CD39-L1, CD39-L2, CD39-L3, and CD39-L4). The family can be subdivided into two groups by conservation of proposed structural domains. The CD39, CD39-L1, and CD39-L3 genes all encode hydrophobic portions in their carboxy and amino termini, serving as transmembrane domains for CD39 and potentially for the other two members. CD39-L2 and CD39-L4 genes encode hydrophobic portions in their amino termini, suggesting that they might encode secreted apyrases. We demonstrate that the CD39-L4 gene encodes the first reported human secreted ecto-apyrase. COS-7 cells transfected with a CD39-L4 expression construct utilizing the naturally occurring leader peptide express recombinant protein outside of the cells. This expression can be blocked by brefeldin A, a chemical that inhibits a step in mammalian secretory pathways. We also demonstrate expression of CD39-L4 message in macrophages, suggesting that the protein is present in the circulation. Furthermore, we show that CD39-L4 is an E-type apyrase, is dependent on calcium and magnesium cations, and has high degree of specificity for NDPs over NTPs as enzymatic substrates. A potential physiological role in hemostasis and platelet aggregation is presented.
人类胞外ATP双磷酸酶基因家族由五个已报道的成员组成（CD39、CD39-L1、CD39-L2、CD39-L3和CD39-L4）。根据推测的结构域保守性，该家族可分为两组。CD39、CD39-L1和CD39-L3基因在其羧基末端和氨基末端均编码疏水部分，作为CD39以及另外两个成员可能的跨膜结构域。CD39-L2和CD39-L4基因在其氨基末端编码疏水部分，这表明它们可能编码分泌型ATP双磷酸酶。我们证明，CD39-L4基因编码首个报道的人类分泌型胞外ATP双磷酸酶。利用天然存在的前导肽转染CD39-L4表达构建体的COS-7细胞在细胞外表达重组蛋白。这种表达可被布雷菲德菌素A阻断，布雷菲德菌素A是一种抑制哺乳动物分泌途径中某一步骤的化学物质。我们还证明了巨噬细胞中CD39-L4信使RNA的表达，这表明该蛋白存在于循环系统中。此外，我们表明CD39-L4是一种E型ATP双磷酸酶，依赖于钙和镁阳离子，并且作为酶底物，对二磷酸核苷（NDPs）的特异性高于三磷酸核苷（NTPs）。文中提出了其在止血和血小板聚集中的潜在生理作用。
