Catalogo Articoli (Spogli Riviste)

OPAC HELP

Titolo:
Histidine 450 plays a critical role in catalysis and, with Ca2+, contributes to the substrate specificity of aminopeptidase A
Autore:
Iturrioz, X; Vazeux, G; Celerier, JM; Corvol, P; Llorens-Cortes, C;
Indirizzi:
Coll France, INSERM Unite 36, F-75005 Paris, France Coll France Paris France F-75005 INSERM Unite 36, F-75005 Paris, France
Titolo Testata:
BIOCHEMISTRY
fascicolo: 11, volume: 39, anno: 2000,
pagine: 3061 - 3068
SICI:
0006-2960(20000321)39:11<3061:H4PACR>2.0.ZU;2-S
Fonte:
ISI
Lingua:
ENG
Soggetto:
STREPTOMYCES-GRISEUS AMINOPEPTIDASE; AMINO-ACID SEQUENCE; ION AFFINITY-CHROMATOGRAPHY; MOLECULAR-CLONING; ANGIOTENSIN-II; METALLOPEPTIDASE FAMILY; ZINC-METALLOPEPTIDASE; BP-1/6C3 ANTIGEN; TAGGED PROTEIN; BINDING-SITE;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
57
Recensione:
Indirizzi per estratti:
Indirizzo: Llorens-Cortes, C Coll France, INSERM Unite 36, 3 Rue Ulm, F-75005 Paris, France Coll France 3 Rue Ulm Paris France F-75005 aris, France
Citazione:
X. Iturrioz et al., "Histidine 450 plays a critical role in catalysis and, with Ca2+, contributes to the substrate specificity of aminopeptidase A", BIOCHEM, 39(11), 2000, pp. 3061-3068

Abstract

Aminopeptidase A (EC 3.4.11.7, APA) is a 130 kDa membrane-bound protease that contains the HEXXH consensus sequence found in the zinc metalloproteasefamily, the zincins. In addition to the catalytic zinc atom, APA contains a Ca2+ ion that increases its enzymatic activity. Aligning the sequences ofthe mouse APA, APN, and other monozinc aminopeptidases led to the identification of a conserved histidine (His 450 in mouse APA). Replacing this residue with a phenylalanine (Phe 450) by site-directed mutagenesis resulted inmarkedly lower levels of APA activity and in a change in the sensitivity of APA to Ca2+ (the EC50 for Ca2+ was 25 mu M in the wild type and only 279 mu M in the mutant). Kinetic studies, with a supramaximal Ca2+ concentration (4 mM), showed that the K-m of the mutant enzyme for the substrate alpha-L-glutamyl-beta-naphthylamide was 25 times higher than that of the wild type, whereas the k(cat) value was much lower (factor of 22). Thus, overall, the wild-type enzyme had a cleavage efficiency that was 571 times higher than that of the mutant. The inhibitory potencies of two different classes of inhibitors, a glutamate thiol and a glutamate phosphonate compound, were significantly lower (factors of 19 and 22, respectively) for the mutated enzyme than for the wild-type enzyme. In contrast, inhibition by lysine thiol was unaffected. These data strongly suggest that His 450 is critical for catalytic activity and is involved in substrate binding via interaction with the P1 carboxylate side chain of the substrate. Furthermore, His 450, together with Ca2+, may contribute to the substrate specificity of APA for N-terminal acidic amino acid residues.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 20/09/20 alle ore 04:18:00