Gene Ontology annotation through association of InterPro records with GO terms
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
Molecular cloning, sequencing, and expression of the gene encoding alkaline ceramidase from Pseudomonas aeruginosa. Cloning of a ceramidase homologue from Mycobacterium tuberculosis.
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The cerN gene was cloned and expressed in E. coli, confirming ceramidase activity.
"Recombinant ceramidase was purified to homogeneity from the lysate of E. coli cells and confirmed to be identical to the Pseudomonas enzyme in its specificity and other enzymatic properties."
Detection of host-derived sphingosine by Pseudomonas aeruginosa is important for survival in the murine lung.
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A clean PA0845 (cerN) deletion abolishes ceramide-induced expression of the sphingosine-regulated sphA; cerN is a member of the SphR regulon.
"we generated a clean deletion in the neutral ceramidase encoded by PA0845 and measured enzyme activity from the PA5325-lacZ reporter construct pAL4. Ceramide fails to induce PA5325-lacZ in the absence of the neutral ceramidase"
Purification and characterization of a novel ceramidase from Pseudomonas aeruginosa.
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Purified P. aeruginosa ceramidase hydrolyses ceramides and catalyses the reverse synthesis; broad substrate range.
"Interestingly, the 70-kDa protein catalyzed a reversible reaction in which the N-acyl linkage of ceramide was either cleaved or synthesized."
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Km 139 uM, Vmax 5.3 umol/min/mg on N-palmitoylsphingosine.
"This bacterial ceramidase had a pH optimum of 8.0-9.0, an apparent Km of 139 microM, and a Vmax of 5.3 micromol/min/mg using N-palmitoylsphingosine as the substrate."