The molecular mechanism of lead inhibition of human porphobilinogen synthase

Human porphobilinogen synthase (PBGS) is a main target in lead poisoning. H uman PBGS purifies with eight Zn(II) per homo-octamer; four ZnA have predom inantly nonsulfur ligands, and four ZnB have predominantly sulfur ligands. Only four Zn(II) are required for activity. To better elucidate the roles o f Zn(TI) and Pb(II), we produced human PBGS mutants that are designed to la ck either the ZnA or ZnB sites. These proteins, MinusZnA (H131A, C223A) and MinusZnB (C122A, C124A, C132A), each become purified with four Zn(II) per octamer, thus confirming an asymmetry in the human PBGS structure. MinusZnA is fully active, whereas MinusZnB is far less active, verifying an importa nt catalytic role for ZnB and the removed cysteine residues. Kinetic proper ties of the mutants and wild type proteins are described, Comparison of Pb( IT) inhibition of the mutants shows that ligands to both ZnA and ZnB intera ct with Pb(II). The ZnB ligands preferentially interact with Pb(II), At lea st one ZnA ligand is responsible for the slow tight binding behavior of Pb( II), The data support a novel model where a high affinity lead site is a hy brid of the ZnA and ZnB sites. We propose that the lone electron pair of Pb (IT) precludes Pb(II) to function in PBGS catalysis.