Purification and characterization of the arylsulfatase synthesized by Pseudomonas aeruginosa PAO during growth in sulfate-free medium and cloning of the arylsulfatase gene (atsA).
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Arylsulfatase purified to homogeneity from PAO1; Km 105 microM for 4-nitrocatechol sulfate; sulfate-repressed; atsA cloned by complementation.
"An arylsulfatase (EC 3.1.6.1) was extracted from Pseudomonas aeruginosa PAO1 and purified 2700-fold to homogeneity"
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AtsA lacks a signal peptide, suggesting a cytoplasmic location.
"The lack of this signal peptide suggests that the P. aeruginosa arylsulfatase is neither periplasmic nor membrane-associated"
Posttranslational formation of formylglycine in prokaryotic sulfatases by modification of either cysteine or serine.
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AtsA carries formylglycine derived from a cysteine; the Cys-to-Ser mutant is inactive.
"Substituting the relevant cysteine by a serine codon in the P. aeruginosa gene led to expression of inactive sulfatase protein, lacking the formylglycine"
1.3 A structure of arylsulfatase from Pseudomonas aeruginosa establishes the catalytic mechanism of sulfate ester cleavage in the sulfatase family.
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Crystal structure shows a formylglycine hydrate as the catalytic residue and Ca2+ in the active site.
"unequivocally showing the presence of a Calpha-formylglycine hydrate as the key catalytic residue"
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The active-site cation is calcium.
"The metal cation involved in stabilizing the charge and anchoring the substrate during catalysis is established as calcium"
Efficient catalytic promiscuity in an enzyme superfamily: an arylsulfatase shows a rate acceleration of 10(13) for phosphate monoester hydrolysis.
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AtsA hydrolyses phosphate monoesters as a promiscuous second activity.
"Besides hydrolyzing sulfate monoesters, this enzyme catalyzes the hydrolysis of phosphate monoesters with multiple turnovers"
Annotation inferences using phylogenetic trees
Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location vocabulary mapping, accompanied by conservative changes to GO terms applied by UniProt
Combined Automated Annotation using Multiple IEA Methods