Human recombinant membrane-bound aminopeptidase P: production of a soluble form and characterization using novel, internally quenched fluorescent substrates

Human recombinant membrane-bound aminopeptidase P: production of a soluble form and characterization using novel, internally quenched fluorescent substrates

Author Molinaro, Giuseppe Google Scholar
Carmona, Adriana Karaoglanovic Autor UNIFESP Google Scholar
Juliano, Maria Aparecida Autor UNIFESP Google Scholar
Juliano, Luiz Autor UNIFESP Google Scholar
Malitskaya, Elena Google Scholar
Yessine, Marie-Andrée Google Scholar
Chagnon, Miguel Google Scholar
Lepage, Yves Google Scholar
Simmons, William H. Google Scholar
Boileau, Guy Google Scholar
Adam, Albert Google Scholar
Institution Univ Montreal
Universidade Federal de São Paulo (UNIFESP)
Loyola Univ
Abstract APP (aminopeptidase P) has the unique ability to cleave the N-terminal amino acid residue from peptides exhibiting a proline at P-1'. Despite its putative involvement in the processing of bioactive peptides, among them the kinins, little is known about the physiological roles of both human forms of APP. The purpose of the present study is first to engineer and characterize a secreted form of hmAPP (human membrane-bound APP). Our biochemical analysis has shown that the expressed glycosylated protein is fully functional, and exhibits enzymic parameters similar to those described previously for mAPP purified from porcine or bovine lungs or expressed from a porcine clone. This soluble form of hmAPP cross-reacts with a polyclonal antiserum raised against a 469-amino-acid hmAPP fragment produced in Escherichia coli. Secondly, we synthesized three internally quenched fluorescent peptide substrates that exhibit a similar affinity for the enzyme than its natural substrates, the kinins, and a higher affinity compared with the tripeptide Arg-Pro-Pro used until now for the quantification of APP in biological samples. These new substrates represent a helpful analytical tool for rapid and reliable: screening of patients susceptible to adverse reactions associated with angiotensin-converting enzyme inhibitors or novel vasopeptidase (mixed angiotensin-converting enzyme/neprilysin) inhibitors.
Keywords adverse reaction
aminopeptidase P
glycosylphosphatidylinositol anchor
internally quenched fluorescent substrate
kinin
screening test
Language English
Date 2005-01-15
Published in Biochemical Journal. London: Portland Press Ltd, v. 385, n. 2, p. 389-397, 2005.
ISSN 0264-6021 (Sherpa/Romeo, impact factor)
Publisher Portland Press Ltd
Extent 389-397
Origin http://dx.doi.org/10.1042/BJ20040849
Access rights Open access Open Access
Type Article
Web of Science ID WOS:000226662500008
URI http://repositorio.unifesp.br/11600/42787

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