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Titolo:
Divalent metal binding properties of the methionyl aminopeptidase from Escherichia coli
Autore:
Dsouza, VM; Bennett, B; Copik, AJ; Holz, RC;
Indirizzi:
Utah State Univ, Dept Chem & Biochem, Logan, UT 84322 USA Utah State UnivLogan UT USA 84322 pt Chem & Biochem, Logan, UT 84322 USA
Titolo Testata:
BIOCHEMISTRY
fascicolo: 13, volume: 39, anno: 2000,
pagine: 3817 - 3826
SICI:
0006-2960(20000404)39:13<3817:DMBPOT>2.0.ZU;2-P
Fonte:
ISI
Lingua:
ENG
Soggetto:
DEPENDENT BETA-LACTAMASE; CYTOCHROME-C-OXIDASE; ACTIVE-SITE; SACCHAROMYCES-CEREVISIAE; AEROMONAS-PROTEOLYTICA; CARBOXYPEPTIDASE-A; CRYSTAL-STRUCTURE; SOLUBLE DOMAIN; SALMONELLA-TYPHIMURIUM; EUKARYOTIC CELLS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
62
Recensione:
Indirizzi per estratti:
Indirizzo: Holz, RC Utah State Univ, Dept Chem & Biochem, Logan, UT 84322 USA Utah State Univ Logan UT USA 84322 Biochem, Logan, UT 84322 USA
Citazione:
V.M. D'souza et al., "Divalent metal binding properties of the methionyl aminopeptidase from Escherichia coli", BIOCHEM, 39(13), 2000, pp. 3817-3826

Abstract

The metal-binding properties of the methionyl aminopeptidase from Escherichia coli (MetAP) were investigated. Measurements of catalytic activity as afunction of added Co(LI) and Fe(II) revealed that maximal enzymatic activity is observed after the addition of only 1 equiv of divalent metal ion. Based on these studies, metal binding constants for the first metal binding event were found to be 0.3 +/- 0.2 mu M and 0.2 +/- 0.2 mu M for Co(II)- andFe(II)-substituted MetAP, respectively. Binding of excess metal ions (>50 equiv) resulted in the loss of similar to 50% of the catalytic activity. Electronic absorption spectral titration of a 1 mM sample of MetAP with Co(II) provided a binding constant of 2.5 +/- 0.5 mM for the second metal binding site. Furthermore, the electronic absorption spectra of Co(II)-loaded MetAP indicated that both metal ions reside in a pentacoordinate geometry. Consistent with the absorption data, electron paramagnetic resonance (EPR) spectra of [CoCo(MetAP)] also indicated that the Co(II) geometries are not highly constrained, suggesting that each Co(II) ion in MetAP resides in a pentacoordinate geometry. EPR studies on [CoCo(MetAP)] also revealed that at pH7.5 there is no significant spin-coupling between the two Co(II), ions, though a small proportion (similar to 5%) Of the sample exhibited detectable spin-spin interactions at pH values > 9.6. EPR studies on [Fe(III)_(MetAP)Iand [Fe(III)Fe(III)(MetAP)1 also suggested no spin-coupling between the two metal ions. H-1 nuclear magnetic resonance (NMR) spectra of [Co(II)_(MetAP)] in both H2O and D2O buffer indicated that the first metal binding site contains the only active-site histidine residue, His171. Mechanistic implications of the observed binding properties of divalent metal ions to the MetAP from E. coli are discussed.

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Documento generato il 01/12/20 alle ore 12:43:37