Characterization of endopeptidase activity of tripeptidyl peptidase-I/CLN2protein which is deficient in classical late infantile neuronal ceroid lipofuscinosis
J. Ezaki et al., Characterization of endopeptidase activity of tripeptidyl peptidase-I/CLN2protein which is deficient in classical late infantile neuronal ceroid lipofuscinosis, BIOC BIOP R, 268(3), 2000, pp. 904-908
Citations number
23
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Endopeptidase activities of the CLN2 gene product (Cln2p)/tripeptidyl pepti
dase I (TPP-I), purified from rat spleen, were studied using the synthetic
fluorogenic substrates. We designed and constructed decapeptides, based on
the known sequence cleavage specificities of bacterial pepstatin-insensitiv
e carboxyl proteases (BPICP) MOCAc-Gly-Lys-Pro-Ile-Pro-Phe-Phe-Arg-Leu-Lys(
Dnp)r-NH2 is readily hydrolyzed by Cln2p/TPP-I (K-cat/K-m = 7.8 s(-1) mM(-1
)). The enzyme had a maximal activity at pH 3.0 for an endopeptidase substr
ate, but at pH 4.5 with respect to tripeptidyl peptidase activity. Both end
opeptidase and tripeptidyl peptidase activities were strongly inhibited by
Ala-Ala-Phe-CH2Cl,, but not inhibited by tyrostatin, an inhibitor of bacter
ial pepstatin-insensitive carboxyl proteases, pepstatin, or inhibitors of s
erine proteases. Fibroblasts from classical late infantile neuronal ceroid
lipofuscinosis patients have less than 5% of the normal tripeptidyl peptida
se activity and pepstatin-insensitive endopeptidase activity. CLn2p/TPP-I i
s a unique enzyme with both tripeptidyl peptidase and endopeptidase activit
ies for certain substrate Specificity. (C) 2000 Academic Press.