GTP-dependent activation of urease apoprotein in complex with the UreD, UreF, and UreG accessory proteins.
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Soriano A, Hausinger RP
GTP-dependent activation of urease apoprotein in complex with the UreD, UreF, and UreG accessory proteins.
Proc Natl Acad Sci U S A. 1999 Sep 28;96(20):11140-4.
- PubMed ID
- 10500143 [ View in PubMed]
- Abstract
Syntheses of metal-containing enzymes often require the participation of accessory proteins. The roles played by many of these accessory proteins are poorly characterized. Klebsiella aerogenes urease, a nickel-containing enzyme, provides an ideal system to study metallocenter assembly. Here, we describe a method for isolating a complex containing urease apoprotein and the UreD, UreF, and UreG accessory proteins. We demonstrate that urease apoprotein in this complex is activated to near wild-type enzyme levels when incubated with nickel ions and high (approximately 100 mM) concentrations of bicarbonate. Significantly, we also observed nickel-dependent activation at physiologically relevant (approximately 100 microM) bicarbonate levels, but only in the presence of GTP. Based on studies involving a nonhydrolyzable analog of GTP, we conclude that nucleotide hydrolysis, not just binding, is required for this process. The critical nucleotide-binding site was localized to UreG on the basis of experiments using a variant complex. These studies highlight the relevance of the UreD-UreF-UreG-urease apoprotein complex to nickel metallocenter assembly and explain the previously identified in vivo energy requirement for urease activation.