Site-directed mutagenesis of two highly conserved residues near the active site of phosphofructo-1-kinase.
Mutations of either of two highly conserved residues near the active site of Escherichia coli phosphofructokinase, Ile-126 or Asn-128, produce no changes in the Km for ATP, relatively small changes in kcat, and a large increase in the Km for fructose 6-P, despite the fact that these residues are not directly involved in substrate binding. A computer graphics analysis of the three-dimensional structure suggests that the mutations effect the orientation of Arg-252, a residue important both for fructose 6-P binding and for catalysis.
Zheng
R L
RL
Department of Biological Chemistry, Chicago Medical School, IL 60064.
Kemp
R G
RG
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D003196:Computer Graphics; D017124:Conserved Sequence; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D010732:Phosphofructokinase-1 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D010732:Phosphofructokinase-1