Identification of oxidized protein hydrolase of human erythrocytes as acylpeptide hydrolase

Citation
T. Fujino et al., Identification of oxidized protein hydrolase of human erythrocytes as acylpeptide hydrolase, BBA-PROT ST, 1478(1), 2000, pp. 102-112
Citations number
30
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY
ISSN journal
0167-4838 → ACNP
Volume
1478
Issue
1
Year of publication
2000
Pages
102 - 112
Database
ISI
SICI code
0167-4838(20000316)1478:1<102:IOOPHO>2.0.ZU;2-K
Abstract
Partial amino acid sequence of 80 kDa oxidized protein hydrolase (OPH), a s erine protease present in human erythrocyte cytosol (Fujino et al., J. Bioc hem. 124 (1998) 1077-1085) that is adherent to oxidized erythrocyte membran es and preferentially degrades oxidatively damaged proteins (Beppu et al., Biochim. Biophys. Acta 1196 (1994) 81-87; Fujino et al., Biochim. Biophys. Acta 1374 (1998) 47-55) was determined. The N-terminal amino acid of diisop ropyl fluorophosphate (DFP)-labeled OPH was suggested to be masked. Six pep tide fragments of OPH obtained by digestion of DFP-labeled OPH with lysyl e ndopeptidase were isolated by use of reverse-phase high-performance liquid chromatography, and the sequence of more than eight amino acids from the N- terminal position of each peptide was determined. Results of homology searc h of amino acid sequence of each peptide strongly suggested that the protei n was identical with human liver acylpeptide hydrolase (ACPH). OPH showed A CPH activity when N-acetyl-L-alanine p-nitroanilide and N-acetylmethionyl L -alanine were used as substrates, Glutathione S-transferase (GST)-tagged re combinant ACPH (rACPH) was prepared by use of baculovirus expression system as a 107-kDa protein from cDNA of human erythroleukemic cell line K-562, r ACPH reacted with anti-OPH antiserum from rabbit. rACPH showed OPH activity when hydrogen Deroxide-oxidized or glycated bovine serum albumin was used as substrates. As well as the enzyme activities of OPH, those of rACPH were inhibited by DFP. The results clearly demonstrate that ACPH, whose physiol ogical function has not yet been well characterized, can play an important role as OPH in destroying oxidatively damaged proteins in living cells. (C) 2000 Elsevier Science B.V. All rights reserved.