Probing the potential metal binding site in Escherichia coli 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (phenylalanine-sensitive).
The active site residues of the proposed metal binding site of DAH 7-P synthase (phe) were probed by site-directed mutagenesis of C61 to glycine and serine, H64 to glycine, and with the double mutant C61H/H64C. While C61S and C61H/ H64C were inactive, both C61G and H64G were active. All mutants, regardless of enzymatic activity, bound one equivalent of Fe2+ per monomeric unit. Even though C61 and H64 were shown not to be metal ligands for the DAH 7-P synthase (phe), they may provide some of the backbone interactions/secondary structural elements necessary to properly form the metal binding pocket.
Sundaram
A K
AK
Interdepartmental Program in Medicinal Chemistry, College of Pharmacy, University of Michigan, Ann Arbor 48109-1065, USA.
Howe
D L
DL
Sheflyan
G Y
GY
Woodard
R W
RW
D010723:3-Deoxy-7-Phosphoheptulonate Synthase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008670:Metals / Q000378:metabolism*; D015335:Molecular Probes; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000494:pharmacology
D017931:DNA Primers; D008670:Metals; D015335:Molecular Probes; D010649:Phenylalanine; D010723:3-Deoxy-7-Phosphoheptulonate Synthase