A broad spectrum Kunitz type serine protease inhibitor secreted by the hookworm Ancylostoma ceylanicum

Aaron Milstone, Lisa M. Harrison, Richard D. Bungiro, Petr Kuzmič, Michael Cappello

Research output: Contribution to journalArticle

Abstract

Although blood-feeding hookworms infect over a billion people worldwide, little is known about the molecular mechanisms through which these parasitic nematodes cause gastrointestinal hemorrhage and iron deficiency anemia. A cDNA corresponding to a secreted Kunitz type serine protease inhibitor has been cloned from adult Ancylostoma ceylanicum hookworm RNA. The translated sequence of the A. ceylanicum Kunitz type inhibitor 1 (AceKI-1) cDNA predicts a 16-amino acid secretory signal sequence, followed by a 68-amino acid mature protein with a molecular mass of 7889 daltons. Recombinant protein (rAceKI-1) was purified from induced lysates of Escherichia coli transformed with the rAceKI-1/pET 28a plasmid, and in vitro studies demonstrate that rAceKI-1 is a tight binding inhibitor of the serine proteases chymotrypsin, pancreatic elastase, neutrophil elastase, and trypsin. AceKI-1 inhibitory activity is present in soluble protein extracts and excretory/secretory products of adult hookworms but not the infective third stage larvae. The native AceKI-1 inhibitor has been purified to homogeneity from soluble extracts of adult A. ceylanicum using size exclusion and reverse-phase high pressure liquid chromatography. As a potent inhibitor of mammalian intestinal proteases, AceKI-1 may play a role in parasite survival and the pathogenesis of hookworm anemia.

Original languageEnglish (US)
Pages (from-to)29391-29399
Number of pages9
JournalJournal of Biological Chemistry
Volume275
Issue number38
DOIs
StatePublished - Sep 22 2000
Externally publishedYes

Fingerprint

Ancylostoma
Ancylostomatoidea
Serine Proteinase Inhibitors
Complementary DNA
High pressure liquid chromatography
Amino Acids
Leukocyte Elastase
Pancreatic Elastase
Chymotrypsin
Molecular mass
Protein Sorting Signals
Recombinant Proteins
Trypsin
Escherichia coli
Proteins
Plasmids
Blood
Peptide Hydrolases
Iron
RNA

ASJC Scopus subject areas

  • Biochemistry

Cite this

A broad spectrum Kunitz type serine protease inhibitor secreted by the hookworm Ancylostoma ceylanicum. / Milstone, Aaron; Harrison, Lisa M.; Bungiro, Richard D.; Kuzmič, Petr; Cappello, Michael.

In: Journal of Biological Chemistry, Vol. 275, No. 38, 22.09.2000, p. 29391-29399.

Research output: Contribution to journalArticle

Milstone, Aaron ; Harrison, Lisa M. ; Bungiro, Richard D. ; Kuzmič, Petr ; Cappello, Michael. / A broad spectrum Kunitz type serine protease inhibitor secreted by the hookworm Ancylostoma ceylanicum. In: Journal of Biological Chemistry. 2000 ; Vol. 275, No. 38. pp. 29391-29399.
@article{178b3dda782c4dfcb08e6c62aba839ca,
title = "A broad spectrum Kunitz type serine protease inhibitor secreted by the hookworm Ancylostoma ceylanicum",
abstract = "Although blood-feeding hookworms infect over a billion people worldwide, little is known about the molecular mechanisms through which these parasitic nematodes cause gastrointestinal hemorrhage and iron deficiency anemia. A cDNA corresponding to a secreted Kunitz type serine protease inhibitor has been cloned from adult Ancylostoma ceylanicum hookworm RNA. The translated sequence of the A. ceylanicum Kunitz type inhibitor 1 (AceKI-1) cDNA predicts a 16-amino acid secretory signal sequence, followed by a 68-amino acid mature protein with a molecular mass of 7889 daltons. Recombinant protein (rAceKI-1) was purified from induced lysates of Escherichia coli transformed with the rAceKI-1/pET 28a plasmid, and in vitro studies demonstrate that rAceKI-1 is a tight binding inhibitor of the serine proteases chymotrypsin, pancreatic elastase, neutrophil elastase, and trypsin. AceKI-1 inhibitory activity is present in soluble protein extracts and excretory/secretory products of adult hookworms but not the infective third stage larvae. The native AceKI-1 inhibitor has been purified to homogeneity from soluble extracts of adult A. ceylanicum using size exclusion and reverse-phase high pressure liquid chromatography. As a potent inhibitor of mammalian intestinal proteases, AceKI-1 may play a role in parasite survival and the pathogenesis of hookworm anemia.",
author = "Aaron Milstone and Harrison, {Lisa M.} and Bungiro, {Richard D.} and Petr Kuzmič and Michael Cappello",
year = "2000",
month = "9",
day = "22",
doi = "10.1074/jbc.M002715200",
language = "English (US)",
volume = "275",
pages = "29391--29399",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "38",

}

TY - JOUR

T1 - A broad spectrum Kunitz type serine protease inhibitor secreted by the hookworm Ancylostoma ceylanicum

AU - Milstone, Aaron

AU - Harrison, Lisa M.

AU - Bungiro, Richard D.

AU - Kuzmič, Petr

AU - Cappello, Michael

PY - 2000/9/22

Y1 - 2000/9/22

N2 - Although blood-feeding hookworms infect over a billion people worldwide, little is known about the molecular mechanisms through which these parasitic nematodes cause gastrointestinal hemorrhage and iron deficiency anemia. A cDNA corresponding to a secreted Kunitz type serine protease inhibitor has been cloned from adult Ancylostoma ceylanicum hookworm RNA. The translated sequence of the A. ceylanicum Kunitz type inhibitor 1 (AceKI-1) cDNA predicts a 16-amino acid secretory signal sequence, followed by a 68-amino acid mature protein with a molecular mass of 7889 daltons. Recombinant protein (rAceKI-1) was purified from induced lysates of Escherichia coli transformed with the rAceKI-1/pET 28a plasmid, and in vitro studies demonstrate that rAceKI-1 is a tight binding inhibitor of the serine proteases chymotrypsin, pancreatic elastase, neutrophil elastase, and trypsin. AceKI-1 inhibitory activity is present in soluble protein extracts and excretory/secretory products of adult hookworms but not the infective third stage larvae. The native AceKI-1 inhibitor has been purified to homogeneity from soluble extracts of adult A. ceylanicum using size exclusion and reverse-phase high pressure liquid chromatography. As a potent inhibitor of mammalian intestinal proteases, AceKI-1 may play a role in parasite survival and the pathogenesis of hookworm anemia.

AB - Although blood-feeding hookworms infect over a billion people worldwide, little is known about the molecular mechanisms through which these parasitic nematodes cause gastrointestinal hemorrhage and iron deficiency anemia. A cDNA corresponding to a secreted Kunitz type serine protease inhibitor has been cloned from adult Ancylostoma ceylanicum hookworm RNA. The translated sequence of the A. ceylanicum Kunitz type inhibitor 1 (AceKI-1) cDNA predicts a 16-amino acid secretory signal sequence, followed by a 68-amino acid mature protein with a molecular mass of 7889 daltons. Recombinant protein (rAceKI-1) was purified from induced lysates of Escherichia coli transformed with the rAceKI-1/pET 28a plasmid, and in vitro studies demonstrate that rAceKI-1 is a tight binding inhibitor of the serine proteases chymotrypsin, pancreatic elastase, neutrophil elastase, and trypsin. AceKI-1 inhibitory activity is present in soluble protein extracts and excretory/secretory products of adult hookworms but not the infective third stage larvae. The native AceKI-1 inhibitor has been purified to homogeneity from soluble extracts of adult A. ceylanicum using size exclusion and reverse-phase high pressure liquid chromatography. As a potent inhibitor of mammalian intestinal proteases, AceKI-1 may play a role in parasite survival and the pathogenesis of hookworm anemia.

UR - http://www.scopus.com/inward/record.url?scp=0034703067&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034703067&partnerID=8YFLogxK

U2 - 10.1074/jbc.M002715200

DO - 10.1074/jbc.M002715200

M3 - Article

C2 - 10893410

AN - SCOPUS:0034703067

VL - 275

SP - 29391

EP - 29399

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 38

ER -