Expression and characterization of elastase inhibitors from the ascarid nematodes Anisakis simplex and Ascaris suum

Tien T. Nguyen, M. A. Qasim, Stephen Morris, Cheng Chan Lu, Dolores Hill, Michael Laskowski, Judy A. Sakanari

研究成果: Article

17 引文 斯高帕斯(Scopus)

摘要

Two elastase inhibitors, ASPI-1 and ASPI-2, from the parasitic nematode Anisakis simplex, have been isolated and characterized. Because these inhibitors are similar in size (60 amino acids in length) and primary sequence (52 and 47% identical) to the Ascaris suum chymotrypsin/elastase inhibitor-1 (AsC/E-1), we suggest that these Anisakis elastase inhibitors belong to the same unique class of canonical inhibitors formed by the family of Ascaris inhibitors (Huang K, Strynadka NCJ, Bernard VD, Peanasky RJ, James MG. Structure 1994;2:679-689). To compare ASPI-1 with AsC/E-1, we expressed both inhibitors in Pichia pastoris and found that: (1) the association constant of rASPI-1 with porcine pancreatic elastase (PPE) is similar to native inhibitor (K(a)=4.5x109 and 6.4x109 M-1, respectively); (2) rASPI-1 is a potent inhibitor of PPE and human leukocyte elastase (K(a)=1.6x109 M-1); and (3) it is only a very weak inhibitor of chymotrypsin (CHYM) (K(a)=1.2x106 M-1). In contrast to the Anisakis inhibitor, however, rAsC/E inhibitor-1 is a very strong inhibitor of both PPE (K(a)=3.5x1010 M-1) and CHYM (K(a)=3.6x1012 M-1). We also found that the determined reactive sites (P1-P1') of rASPI-1 and rAsC/E-1, as recognized by PPE, are Ala 28-Met 29 and Leu 31-Met 32, respectively. These P1-P1' residues of AsC/E-1 constitute the same reactive site as that also recognized by CHYM (Peanasky RJ, Bentz Y, Homandberg GA, Minor ST, Babin DR. Arch Biochem Biophys 1994;232:135-142). The difference in specificities of ASPI-1 and AsC/E-1 toward their cognate serine proteases may be attributed to the P1 and P3' residues in the inhibitors. Elastase, which recognizes both alanine and leucine, can accommodate both ascarid inhibitors, whereas chymotrypsin, which prefers bulky, hydrophobic residues, only recognizes the Ascaris C/E inhibitor-1. Copyright (C) 1999 Elsevier Science B.V.

原文English
頁(從 - 到)79-89
頁數11
期刊Molecular and Biochemical Parasitology
102
發行號1
DOIs
出版狀態Published - 1999 七月 30

All Science Journal Classification (ASJC) codes

  • Parasitology
  • Molecular Biology

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