Kinetics and thermodynamics of the interchange of the morpheein forms of human porphobilinogen synthase.
A morpheein is a homo-oligomeric protein that can adopt different nonadditive quaternary assemblies (morpheein forms) with different functionalities. The human porphobilinogen synthase (PBGS) morpheein forms are a high activity octamer, a low activity hexamer, and two structurally distinct dimer conformations. Conversion between hexamer and octamer involves dissociation to dimers, conformational change at the dimer level, followed by association to the alternate assembly. The current work promotes an alternative and novel view of the physiologically relevant dimeric structures, which are derived from the crystal structures, but are distinct from the asymmetric units of their crystal forms. Using a well characterized heteromeric system (WT+F12L; Tang, L. et al. (2005) J. Biol. Chem. 280, 15786-15793), extensive study of the human PBGS morpheein reequilibration process now reveals that the intervening dimers do not dissociate to monomers. The morpheein equilibria of wild type (WT) human PBGS are found to respond to changes in pH, PBGS concentration, and substrate turnover. Notably, the WT enzyme is predominantly an octamer at neutral pH, but increasing pH results in substantial conversion to lower order oligomers. Most significantly, the free energy of activation for the conversion of WT+F12L human PBGS heterohexamers to hetero-octamers is determined to be the same as that for the catalytic conversion of substrate to product by the octamer, remarkably suggesting a common rate-limiting step for both processes, which is postulated to be the opening/closing of the active site lid.
Selwood
Trevor
T
Fox Chase Cancer Center, 333 Cottman Avenue, Philadelphia, Pennsylvania 19111, USA.
Tang
Lei
L
Lawrence
Sarah H
SH
Anokhina
Yana
Y
Jaffe
Eileen K
EK
D019281:Dimerization; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D000623:Porphobilinogen Synthase / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013816:Thermodynamics
D000623:Porphobilinogen Synthase