p-Aminobenzoate synthesis in Escherichia coli: purification and characterization of PabB as aminodeoxychorismate synthase and enzyme X as aminodeoxychorismate lyase.
The Escherichia coli pabA and pabB genes have been overexpressed separately and in tandem. Using purified PabB, we have confirmed the recent suggestion that PabB needs an additional protein, enzyme X, to convert chorismate and NH3 to p-aminobenzoate (PABA). With chorismate and NH3, pure PabB generates an intermediate presumed to be 4-amino-4-deoxychorismate based upon UV/visible spectroscopy and enzymatic and nonenzymatic transformations. The PabB-catalyzed interconversion of chorismate and isolated aminodeoxychorismate is readily reversible. With pure PabB as a stoichiometric assay reagent, enzyme X was purified approximately 800-fold to near homogeneity as an apparent homodimer of 50 kDa from E. coli. Enzyme X shows no activity on chorismate but quantitatively converts the preformed aminodeoxychorismate into p-aminobenzoate and pyruvate, acting thereby as an aminodeoxychorismate lyase.
Ye
Q Z
QZ
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115.
Liu
J
J
Walsh
C T
CT
D010129:4-Aminobenzoic Acid / Q000378:metabolism*; D001483:Base Sequence; D019731:Carbon-Nitrogen Ligases; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005798:Genes, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D007652:Oxo-Acid-Lyases / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D000637:Transaminases / Q000235:genetics* / Q000378:metabolism
D015345:Oligonucleotide Probes; D000637:Transaminases; C066599:aminodeoxychorismate synthase; D007652:Oxo-Acid-Lyases; C066600:aminodeoxychorismate lyase; D019731:Carbon-Nitrogen Ligases; D010129:4-Aminobenzoic Acid