Molecular cloning, chromosomal localization, and bacterial expression of a murine macrophage metalloelastase.
Molecular cloning, chromosomal localization, and bacterial expression of a murine macrophage metalloelastase.
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DOI：
10.1016/s0021-9258(18)42885-2
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发表时间：
1992-03
期刊：
影响因子：
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通讯作者：
S. Shapiro;G. Griffin;D. Gilbert;N. Jenkins;N. Copeland;H. Welgus;R. Senior;T. Ley
中科院分区：
文献类型：
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作者：
S. Shapiro;G. Griffin;D. Gilbert;N. Jenkins;N. Copeland;H. Welgus;R. Senior;T. Ley
此前已有研究表明，小鼠巨噬细胞可以分泌一种锌依赖的蛋白水解酶，这种酶可以降解弹性蛋白。在这篇报道中，我们鉴定了小鼠巨噬细胞弹性蛋白酶(MME)基因，并证明它是金属蛋白酶基因家族中的一个独特的成员。对少量MME进行纯化，得到N-末端氨基酸序列。利用该序列通过聚合酶链式反应获得部分c DNA克隆，并用该探针筛选小鼠巨噬细胞样细胞系P388D 1的c DNA文库。全长约1.8kb的MME基因包含一个1386个碱基对的开放阅读框，预测的MME酶原分子量为53 kDa。编码Mme的基因在小鼠基因组中只表达一次，位于9号染色体上。尽管Mme的大小与其他金属蛋白酶相似，但Mme却是不同的，与其他金属蛋白酶的氨基酸同源性只有33-48%。与其他金属酶不同的是，MME似乎被快速加工成活性截断形式(N-端和C-端切割)。我们在大肠杆菌中表达了重组MME，并证明其具有显著的弹性溶解活性，该活性可被金属蛋白酶组织抑制物特异性地抑制。因此，MME是一种真正的金属蛋白酶，可能参与组织损伤和重塑。
Murine macrophages have previously been shown to secrete a zinc-dependent proteinase that can degrade elastin. In this report, we identify murine macrophage elastase (MME) cDNA and show that it is a distinct member of the metalloproteinase gene family. Small amounts of MME were purified to homogeneity, and N-terminal amino acid sequence was obtained. This sequence was used to obtain a partial cDNA clone by the polymerase chain reaction; a cDNA library derived from a mouse macrophage-like cell line (P388D1) was screened with this probe. A full-length MME cDNA spanning approximately 1.8 kilobases contained an open reading frame of 1386 base pairs; the predicted molecular mass of the MME proenzyme is 53 kDa. The gene encoding MME is represented only once in the mouse genome and is located on chromosome 9. Despite a size that is similar to other metalloproteinases, MME is distinct, sharing only 33-48% amino acid homology with other metalloproteinases. In contrast to other metalloenzymes, MME appears to be rapidly processed to an active truncated form (N-terminal and C-terminal cleavage). We expressed recombinant MME in Escherichia coli and demonstrated that it has significant elastolytic activity that is specifically inhibited by the tissue inhibitor of metalloproteinases. MME is therefore a true metalloproteinase that may be involved in tissue injury and remodeling.