Cloning of the gene and characterization of the enzymatic properties of the monomeric alkaline phosphatase (PhoX) from Pasteurella multocida strain X-73.
We have identified a new phoX gene encoding the monomeric alkaline phosphatase from Pasteurella multocida X-73. This gene was not found in the published genome sequence of Pasteurella multocida pm70. Characterization of the recombinant PhoX of Pasteurella multocida X-73 showed that it is a monomeric enzyme, activated by Ca(2+) and possibly secreted by the Tat pathway. These features distinguish phosphatases of the PhoX family from those of the PhoA family. All proteins of the PhoX family were found to contain a conserved motif that shares significant sequence homology with the calcium-binding site of a phosphotriesterase known as diisopropylfluorophosphatase. Site-directed mutagenesis revealed that D527 of PhoX might be the ligand bound to the catalytic calcium. This is the first report on identification of homologous sequences between PhoX and the phosphotriesterase and on the potential calcium-binding site of PhoX.
Wu
Jin-Ru
JR
Graduate Institute of Veterinary Microbiology, National Chung Hsing University, Taichung, Taiwan.
Shien
Jui-Hung
JH
Shieh
Happy K
HK
Hu
Chung-Chi
CC
Gong
Shuen-Rong
SR
Chen
Ling-Yun
LY
Chang
Poa-Chun
PC
D000469:Alkaline Phosphatase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002118:Calcium / Q000494:pharmacology; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D020536:Enzyme Activators / Q000494:pharmacology; D008024:Ligands; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016979:Pasteurella multocida / Q000201:enzymology* / Q000235:genetics*; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000378:metabolism; D017422:Sequence Analysis, DNA
D004269:DNA, Bacterial; D020536:Enzyme Activators; D008024:Ligands; D021122:Protein Subunits; D011994:Recombinant Proteins; D000469:Alkaline Phosphatase; D002118:Calcium