Building reactive copper centers in human carbonic anhydrase II.
J Biol Inorg Chem
; 18(6): 595-8, 2013 Aug.
Article
em En
| MEDLINE
| ID: mdl-23744511
ABSTRACT
Reengineering metalloproteins to generate new biologically relevant metal centers is an effective a way to test our understanding of the structural and mechanistic features that steer chemical transformations in biological systems. Here, we report thermodynamic data characterizing the formation of two type-2 copper sites in carbonic anhydrase and experimental evidence showing one of these new, copper centers has characteristics similar to a variety of well-characterized copper centers in synthetic models and enzymatic systems. Human carbonic anhydrase II is known to bind two Cu(2+) ions; these binding events were explored using modern isothermal titration calorimetry techniques that have become a proven method to accurately measure metal-binding thermodynamic parameters. The two Cu(2+)-binding events have different affinities (K a approximately 5 × 10(12) and 1 × 10(10)), and both are enthalpically driven processes. Reconstituting these Cu(2+) sites under a range of conditions has allowed us to assign the Cu(2+)-binding event to the three-histidine, native, metal-binding site. Our initial efforts to characterize these Cu(2+) sites have yielded data that show distinctive (and noncoupled) EPR signals associated with each copper-binding site and that this reconstituted enzyme can activate hydrogen peroxide to catalyze the oxidation of 2-aminophenol.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Compostos Organometálicos
/
Cobre
/
Anidrase Carbônica II
Tipo de estudo:
Prognostic_studies
Limite:
Humans
Idioma:
En
Revista:
J Biol Inorg Chem
Assunto da revista:
BIOQUIMICA
Ano de publicação:
2013
Tipo de documento:
Article
País de afiliação:
Estados Unidos