Wiesner Olaf, Litwiller Robert D, Hummel Amber M, Viss Margaret A, McDonald Cari J, Jenne Dieter E, Fass David N, Specks U
Thoracic Diseases Research Unit, Division of Pulmonary and Critical Care Medicine, Mayo Clinic Foundation, Rochester, MN 55905, USA.
FEBS Lett. 2005 Oct 10;579(24):5305-12. doi: 10.1016/j.febslet.2005.08.056.
Direct comparisons of human (h) and murine (m) neutrophil elastase (NE) and proteinase 3 (PR3) are important for the understanding and interpretation of inflammatory and PR3-related autoimmune processes investigated in wild-type-, mNE- and mPR3/mNE knockout mice. To this end, we purified recombinant mPR3 and mNE expressed in HMC1 and 293 cells and compared their biophysical properties, proteolytic activities and susceptibility to inhibitors with those of their human homologues, hPR3 and hNE. Significant species differences in physico-chemical properties, substrate specificities and enzyme kinetics towards synthetic peptide substrates, oxidized insulin B chain, and fibrinogen were detected. MeOSuc-AAPV-pNA and Suc-AAPV-pNA were hydrolyzed more efficiently by mPR3 than hPR3, but enzymatic activities of mNE and hNE were very similar. Fibrinogen was cleaved much more efficiently by mPR3 than by hPR3. All four proteases were inhibited by alpha(1)-antitrypsin and elafin. Eglin C inihibited mNE, hNE, mPR3, but not hPR3. SLPI inhibited both NEs, but neither PR3. The custom-designed hNE inhibitor, Val(15)-aprotinin, is a poor inhibitor for mNE. In conclusion, appropriate interpretation of experiments in murine models requires individual species-specific assessment of neutrophil protease function and inhibition.
对人类（h）和小鼠（m）中性粒细胞弹性蛋白酶（NE）及蛋白酶3（PR3）进行直接比较，对于理解和解释在野生型、mNE和mPR3/mNE基因敲除小鼠中所研究的炎症及PR3相关自身免疫过程至关重要。为此，我们纯化了在HMC1和293细胞中表达的重组mPR3和mNE，并将它们的生物物理特性、蛋白水解活性以及对抑制剂的敏感性与其人类同源物hPR3和hNE进行了比较。在物理化学性质、底物特异性以及针对合成肽底物、氧化胰岛素B链和纤维蛋白原的酶动力学方面，检测到了显著的种属差异。mPR3对MeOSuc - AAPV - pNA和Suc - AAPV - pNA的水解效率高于hPR3，但mNE和hNE的酶活性非常相似。mPR3对纤维蛋白原的切割效率远高于hPR3。所有四种蛋白酶均受到α1 - 抗胰蛋白酶和弹性蛋白酶抑制因子的抑制。艾格林C抑制mNE、hNE和mPR3，但不抑制hPR3。分泌性白细胞蛋白酶抑制因子抑制两种NE，但不抑制任何一种PR3。定制设计的hNE抑制剂Val(15) - 抑肽酶对mNE的抑制效果不佳。总之，对小鼠模型实验进行恰当的解释需要对中性粒细胞蛋白酶的功能和抑制作用进行针对每个物种的特异性评估。
