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 Escherichia coli with its allosteric activator, N-acetyl-D-glucosamine 6-phosphate, was studied by different spectrophotometric methods. Analysis of the circular-dichroism differential spectra produced by the binding of the allosteric activator or the competitive inhibitor 2-amino-2-deoxy-D-glucitol 6-phosphate (a homotropic ligand displacing the allosteric equilibrium to the R conformer), strongly suggests the presence of tyrosine residues at or near the allosteric site, although a conformational effect cannot be ruled out. The involvement of a single tyrosine residue in the N-acetyl-D-glucosamine-6-phosphate binding site of glucosamine-6-phosphate deaminase was supported by spectrophotometric pH titrations performed in the presence or absence of the homotropic and heterotropic ligand. In these experiments, a single titrated tyrosine residue is completely protected by saturation with the allosteric activator; this group is considerably acidic (pK 8.75). The analysis of the amino acid sequence of the deaminase using a set of indices for the prediction of surface accessibility of amino acid residues, suggests that the involved residue may be Tyr121 or Tyr254.
Altamirano
M M
MM
Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, Distrito Federal, Mexico.
Hernandez-Arana
A
A
Tello-Solis
S
S
Calcagno
M L
ML
D019747:Aldose-Ketose Isomerases; D000495:Allosteric Site; D000595:Amino Acid Sequence; D002238:Carbohydrate Epimerases / Q000737:chemistry* / Q000302:isolation & purification; D002942:Circular Dichroism; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D014443:Tyrosine
D014443:Tyrosine; C020582:glucosamine-6-phosphate isomerase; D002238:Carbohydrate Epimerases; D019747:Aldose-Ketose Isomerases