Cloning and characterization of novel mouse and human secretory phospholipase A2s
Cloning and characterization of novel mouse and human secretory phospholipase A2s
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DOI：
10.1074/jbc.274.35.24973
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发表时间：
1999-08-27
影响因子：
4.8
通讯作者：
Hanasaki, K
中科院分区：
文献类型：
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作者：
Ishizaki, J;Suzuki, N;Hanasaki, K
哺乳动物分泌性磷脂酶A(2)s （sPLA(2)s）根据分子结构和分子内二硫桥的定位可分为几类。其中，IIA组sPLA(2)在各种炎症条件下表达增强，被认为是炎症性疾病发病的关键酶之一。然而，在许多近亲繁殖的小鼠品系中，IIA组sPLA(2)基因被移码突变自然破坏。在这里，我们报道了克隆一种编码sPLA(2)在IIA组sPLA(2)缺陷小鼠脾脏中表达的新sPLA(2)的cDNA。我们还克隆了它的人类同源物，并将其基因定位在染色体1p36.12上，靠近IIA和V sPLA(2)基因的位点。人类成熟的sPLA(2)蛋白由125个氨基酸（M-r = 14,500）组成，前面有一个20个残基的前肽，在半胱氨酸残基的数量和位置以及总体特性方面与IIA组sPLA(2)最相似（48%）。基于这些结构特性，新型sPLA(2)应被归类为II类，称为IID类，以遵循已经确定的IIA到IIC sPLA(2)s。当cDNA在COS-7细胞中表达时，PLA(2)活性优先在培养基中积累。它在中性至碱性pH值和2毫米Ca2+时具有最大活性。在对单个底物的分析中，l- α -1-棕榈酰-2-亚油基磷脂酰乙醇胺的水解效率高于所检测的其他磷脂。与cDNA杂交的RNA印迹在人的脾脏、胸腺和结肠中发现了两个转录本（2.0和1.0 kb）。在内毒素处理大鼠和IIA组sPLA(2)缺陷小鼠胸腺中，一种新型sPLA(2) mRNA的表达升高，表明其在炎症过程中的功能作用。
Mammalian secretory phospholipase A(2)s (sPLA(2)s) are classified into several groups according to molecular structure and the localization of intramolecular disulfide bridges. Among them, group IIA sPLA(2) has been thought to be one of the key enzymes in the pathogenesis of inflammatory diseases owing to its augmented expression under various inflammatory conditions. However, in a number of inbred mouse strains, the group IIA sPLA(2) gene is naturally disrupted by a frameshift mutation. Here, we report the cloning of a cDNA encoding a novel sPLA(2) expressed in the spleen of group IIA sPLA(2)-deficient mouse. We also cloned its human homolog and mapped its gene location on chromosome 1p36.12 near the loci of group IIA and V sPLA(2) genes. The human mature sPLA(2) protein consists of 125 amino acids (M-r = 14,500) preceded by a 20-residue prepeptide and is most similar to group IIA sPLA(2) with respect to the number and positions of cysteine residues as well as overall identity (48%). Based on these structural properties, the novel sPLA(2) should be categorized into group II, called group IID to follow the already identified IIA to IIC sPLA(2)s. When the cDNA was expressed in COS-7 cells, PLA(2) activity preferentially accumulated in the culture medium. It is maximally active at neutral to alkaline pH and with 2 mm Ca2+. In assays with individual substrates, L-alpha-1-palmitoyl-2-linoleoyl phosphatidylethanolamine was more efficiently hydrolyzed than the other phospholipids examined. An RNA blot hybridized with the cDNA exhibited two transcripts (2.0 and 1.0 kb) in human spleen, thymus, and colon. The expression of a novel sPLA(2) mRNA was elevated in the thymus after treatment with endotoxin in rats as well as in group IIA sPLA(2)-deficient mice, suggesting its functional role in the progression of the inflammatory process.