The Ala-161-->Thr substitution in Escherichia coli alkaline phosphatase does not result in loss of enzymatic activity although the homologous mutation in humans causes hypophosphatasia.
The correlation between sequence homology and catalytic importance of a specific amino acid in the E. coli and Liver/Kidney/Bone (L/K/B) human alkaline phosphatase was investigated. For this reason Ala-161 in the E. coli enzyme was substituted with a threonine residue via site-specific mutagenesis. The homologous amino acid in the L/K/B alkaline phosphatase sequence has been shown to cause inactivation of the enzyme. In E. coli alkaline phosphatase the Ala-161-->Thr substitution results in a mutant enzyme with virtually unchanged catalytic properties when compared to the wild-type enzyme. Our results show that Ala-161 in the E. coli alkaline phosphatase does not have an important catalytic role. The results suggest that the three-dimensional topology of the L/K/B alkaline phosphatase may be different from that observed for the enzyme from E. coli.
Chaidaroglou
A
A
Department of Chemistry, Merkert Chemistry Center, Boston College, Chestnut Hill, MA 02167.
Kantrowitz
E R
ER
D000409:Alanine; D000469:Alkaline Phosphatase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001842:Bone and Bones / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D006801:Humans; D007014:Hypophosphatasia / Q000201:enzymology*; D007527:Isoenzymes / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013912:Threonine
D007527:Isoenzymes; D011994:Recombinant Proteins; D013912:Threonine; D000469:Alkaline Phosphatase; D000409:Alanine