The
enzyme responsible for the hydrolysis of
phosphonoacetic acid, a non-biogenic C-P compound, was purified to electrophoretic homogeneity from a wild-type strain of Penicillium oxalicum. A 50-fold enrichment was obtained by a combination of
anion exchange, hydrophobic interaction and MonoQ-fast
protein liquid chromatography, with a yield of one-third of the initial activity. A characterization of the
protein showed both similarities and differences with respect to the well-characterized bacterial counterpart. The fungal
phosphonoacetate hydrolase is a 43-kDa monomeric
protein showing low affinity toward its substrate and high sensitivity to even mildly acidic pH values.
Enzyme activity neither required nor was stimulated by the presence of
divalent cations. Polyclonal
antibodies were raised in mouse against the purified
protein, allowing the study of enzyme induction as a function of the
phosphate status of the cell.
Peptide mass mapping led to the determination of about 20% of the primary structure. Despite the biochemical differences,
amino acid alignment showed a high degree of similarity of the fungal
hydrolase with the few sequences available to date for the bacterial
enzyme. The possible physiological role of a
phosphonoacetate hydrolase is discussed.