Conversion of the Alzheimer's beta-amyloid precursor protein (APP) Kunitz domain into a potent human neutrophil elastase inhibitor.

نویسندگان

  • S Sinha
  • J Knops
  • F Esch
  • E D Moyer
  • T Oltersdorf
چکیده

Site-specific mutagenesis techniques have been used to construct active site variants of the Kunitz-type protease inhibitor domain present in the Alzheimer's beta-amyloid precursor protein (APP-KD). Striking alteration of its protease inhibitory properties were obtained when the putative P1 residue, arginine, was replaced with the small hydrophobic residue valine. The altered protein was no longer inhibitory toward bovine pancreatic trypsin, human Factor XIa, mouse epidermal growth factor-binding protein, or bovine chymotrypsin, all of which are strongly inhibited by the unaltered APP-KD (Sinha, S., Dovey, H. F., Seubert, P., Ward, P. J., Blacher, R. W., Blaber, M., Bradshaw, R. A., Arici, M., Mobley, W. C., and Lieberburg, I. (1990) J. Biol. Chem. 265, 8983-8985). Instead, the P1-Val-APP-KD was a potent inhibitor of human neutrophil elastase, with a Ki = 0.8 nM, as estimated by the inhibition of the activity of human neutrophil elastase measured using a chromogenic substrate. It also inhibited the degradation of insoluble elastin by the enzyme virtually stoichiometrically. Replacement of the P1' (Ala) and P2' (Met) residues of P1-Val-MKD with the corresponding residues (Ser, Ile) from alpha 1-proteinase inhibitor resulted in an inactive protein, underscoring the mechanistic differences between the serpins from the Kunitz-type protease inhibitor family. These results confirm the importance of the P1 arginine residue of APP-KD in determining inhibitory specificity, and are also the first time that a single amino acid replacement has been shown to generate a specific potent human neutrophil elastase inhibitor from a human KD sequence.

برای دانلود رایگان متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Potential role of protease nexin-2/amyloid beta-protein precursor as a cerebral anticoagulant.

The amyloid beta-protein precursor (APP) is the parent molecule to the amyloid beta-protein which is a major constituent of neuritic plaques and cerebrovascular deposits in Alzheimer's disease (AD). The protease inhibitor, protease nexin-2 (PN-2), is the secreted form of APP that contains the Kunitz protease inhibitor (KPI) domain. We reported that the predominant isoform of APP in human brain ...

متن کامل

The amyloid precursor protein/protease nexin 2 Kunitz inhibitor domain is a highly specific substrate of mesotrypsin.

The amyloid precursor protein (APP) is a ubiquitously expressed transmembrane adhesion protein and the progenitor of amyloid-beta peptides. The major splice isoforms of APP expressed by most tissues contain a Kunitz protease inhibitor domain; secreted APP containing this domain is also known as protease nexin 2 and potently inhibits serine proteases, including trypsin and coagulation factors. T...

متن کامل

Characterization of platelet-releasable forms of beta-amyloid precursor proteins: the effect of thrombin.

Activated platelets release a potent inhibitor of factor XIa previously identified as a Kunitz proteinase inhibitor domain-containing form of the beta-amyloid precursor proteins (beta APP). Two carboxy-terminal truncated forms of the beta APP, beta APP-751 and beta APP-770, are shown to be the predominant isoforms secreted by platelets. The release of beta APP from platelets is responsible for ...

متن کامل

Demonstration by fluorescence resonance energy transfer of two sites of interaction between the low-density lipoprotein receptor-related protein and the amyloid precursor protein: role of the intracellular adapter protein Fe65.

Amyloid-beta, the major constituent of senile plaques in Alzheimer's disease, is derived from the amyloid precursor protein (APP) by proteolysis. Kunitz protease inhibitor (KPI) containing forms of APP (APP751/770) interact with a multifunctional endocytic receptor, the low-density lipoprotein receptor-related protein (LRP), which modulates its proteolytic processing affecting production of amy...

متن کامل

Post-translational processing and turnover kinetics of presynaptically targeted amyloid precursor superfamily proteins in the central nervous system.

The amyloid precursor superfamily is composed of three highly conserved transmembrane glycoproteins, the amyloid precursor protein (APP) and amyloid precursor-like proteins 1 and 2 (APLP1, APLP2), whose functions are unknown. Proteolytic cleavage of APP yields the betaA4 peptide, the major component of cerebral amyloid in Alzheimer's disease. Here we show that five post-translationally modified...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

عنوان ژورنال:
  • The Journal of biological chemistry

دوره 266 31  شماره 

صفحات  -

تاریخ انتشار 1991