Structures of the Klebsiella aerogenes urease apoenzyme and two active-site mutants

Urease from Klebsiella aerogenes [Jabri et al. (1995) Science 268, 998-1004] is an (alpha beta gamma)(3) trimer with each alpha-subunit having an (alpha beta)(8)-barrel domain containing a binickel active center. Here we examine structure-function relations for urease in more detail through structural analysis of the urease apoenzyme at 2.3 Angstrom resolution and mutants of two key catalytic residues (H219A and H320A) at 2.5 Angstrom resolution. With the exception of the active site, in which a water molecule takes the place of the missing carbamate and nickel atoms, the structure of the apoenzyme is nearly identical to that of the holoenzyme, suggesting a high degree of preorganization which helps explain the tight binding of nickel, In the structure of H219A, the major change involves a conformational shift and ordering of the active site flap, but a small shift in the side chain of Asp(alpha 211) could contribute to the lower activity of H219A. In the H320A structure, the catalytic water, primarily a Ni-2 ligand in the holoenzyme, shifts into a bridging position. This shift shows that the nickel ligation is rather sensitive to the environment and the change in ligation may contribute to the 10(5)-fold lower activity of H320A. In addition, these results show that urease is resilient to the loss of nickel ions and mutations, Analysis of the urease tertiary/quaternary structure suggests that the stability of this enzyme may be largely due to its burial of an unusually large fraction of its residues: 50% in the gamma-subunit, 30% in the beta-subunit, and 60% in the alpha-subunit.