Elafin, an Elastase-specific Inhibitor, Is Cleaved by Its Cognate Enzyme Neutrophil Elastase in Sputum from Individuals with Cystic Fibrosis
Elafin, an Elastase-specific Inhibitor, Is Cleaved by Its Cognate Enzyme Neutrophil Elastase in Sputum from Individuals with Cystic Fibrosis
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DOI：
10.1074/jbc.m803707200
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发表时间：
2008-11-21
影响因子：
4.8
通讯作者：
McElvaney, Noel G.
中科院分区：
文献类型：
--
作者：
Guyot, Nicolas;Butler, Marcus W.;McElvaney, Noel G.
Elafin是一种中性粒细胞丝氨酸蛋白酶抑制剂，在肺中表达，并显示抗炎和抗菌特性。先前的研究表明，囊性纤维化（CF）和慢性阻塞性肺病气道的一些先天宿主防御分子由于在肺部炎症期间观察到的蛋白水解降解增加而受损。鉴于这些发现，我们因此集中于CF肺中弹性蛋白酶的状态。我们在本研究中发现，弹性蛋白酶在CF患者的痰液中被裂解。与铜绿假单胞菌阴性样品相比，发现铜绿假单胞菌阳性CF痰含有较低的弹性蛋白水平和较高的中性粒细胞弹性蛋白酶（NE）活性，其在切割重组弹性蛋白酶方面特别有效。NE在该过程中起着关键作用，因为只有NE抑制剂能够抑制弹性蛋白降解。进一步的体外研究表明，重组弹性蛋白酶与过量NE的孵育导致抑制剂的快速裂解。在弹力蛋白的N-末端鉴定出两个切割位点（瓦尔-5-Lys-6和瓦尔-9-Ser-10）。有趣的是，抑制剂的纯化片段（Lys-6-Gln-57和Ser-10-Gln-57）显示出仍然具有抑制NE的活性。然而，过量的NE显示出强烈降低弹性蛋白酶结合脂多糖（LPS）的能力及其通过转谷氨酰胺化固定的能力。总之，本研究提供的证据表明，弹性蛋白酶被CF痰液中浓度过高的同源酶NE切割，铜绿假单胞菌感染促进了这种作用。这种切割可能对弹性蛋白的先天免疫功能有影响。
Elafin is a neutrophil serine protease inhibitor expressed in lung and displaying anti-inflammatory and anti-bacterial properties. Previous studies demonstrated that some innate host defense molecules of the cystic fibrosis (CF) and chronic obstructive pulmonary disease airways are impaired due to increased proteolytic degradation observed during lung inflammation. In light of these findings, we thus focused on the status of elafin in CF lung. We showed in the present study that elafin is cleaved in sputum from individuals with CF. Pseudomonas aeruginosa-positive CF sputum, which was found to contain lower elafin levels and higher neutrophil elastase (NE) activity compared with P. aeruginosa-negative samples, was particularly effective in cleaving recombinant elafin. NE plays a pivotal role in the process as only NE inhibitors are able to inhibit elafin degradation. Further in vitro studies demonstrated that incubation of recombinant elafin with excess of NE leads to the rapid cleavage of the inhibitor. Two cleavage sites were identified at the N-terminal extremity of elafin (Val-5-Lys-6 and Val-9-Ser-10). Interestingly, purified fragments of the inhibitor (Lys-6-Gln-57 and Ser-10-Gln-57) were shown to still be active for inhibiting NE. However, NE in excess was shown to strongly diminish the ability of elafin to bind lipopolysaccharide(LPS) and its capacity to be immobilized by transglutamination. In conclusion, this study provides evidence that elafin is cleaved by its cognate enzyme NE present at excessive concentration in CF sputum and that P. aeruginosa infection promotes this effect. Such cleavage may have repercussions on the innate immune function of elafin.