SPECTROCHEMICAL EVIDENCE FOR THE PRESENCE OF A TYROSINE RESIDUE IN THE ALLOSTERIC SITE OF GLUCOSAMINE-6-PHOSPHATE DEAMINASE FROM ESCHERICHIA-COLI

The interaction of the enzyme glucosamine 6-phosphate deaminase from E scherichia coli with its allosteric activator, N-acetyl-D-glucosamine 6-phosphate, was studied by different spectrophotometric methods. Anal ysis of the circular-dichroism differential spectra produced by the bi nding of the allosteric activator or the competitive inhibitor 2-amino -2-deoxy-D-glucitol 6-phosphate (a homotropic ligand displacing the al losteric equilibrium to the R conformer), strongly suggests the presen ce of tyrosine residues at or near the allosteric site, although a con formational effect cannot be ruled out. The involvement of a single ty rosine residue in the N-acetyl-D-glucosamine-6-phosphate binding site of glucosamine-6-phosphate deaminase was supported by spectrophotometr ic pH titrations performed in the presence or absence of the homotropi c and heterotropic ligand. In these experiments, a single titrated tyr osine residue is completely protected by saturation with the allosteri c activator; this group is considerably acidic (pK 8.75). The analysis of the amino acid sequence of the deaminase using a set of indices fo r the prediction of surface accessibility of amino acid residues, sugg ests that the involved residue may be Tyr121 or Tyr254.