REQUIREMENT OF CARBON-DIOXIDE FOR IN-VITRO ASSEMBLY OF THE UREASE NICKEL METALLOCENTER

Assembly of protein metallocenters is not well understood. Urease offers a tractable system for examination of this process. Formation of the urease metallocenter in vivo is known to require four accessory proteins: UreD, postulated to be a urease-specific molecular chaperone; UreE, a nickel(II)-binding protein; and UreF and UreG, of unknown function. Activation of purified Klebsiella aerogenes urease apoprotein was accomplished in vitro by providing carbon dioxide (half-maximal activation at similar to 0.2 percent carbon dioxide) in addition to nickel ion. Activation coincided with carbon dioxide incorporation into urease in a pH-dependent reaction (pK(a) greater than or equal to 9, where K-a is the acid constant). The concentration of carbon dioxide also affected the amount of activation of UreD-urease apoprotein complexes. These results suggest that carbon dioxide binding to urease apoprotein generates a ligand that facilitates productive nickel binding.