Literature summary for 3.1.1.3 extracted from

Hausmann, S.; Wilhelm, S.; Jaeger, K.E.; Rosenau, F.
Catalytic mechanism of C-di-GMP specific phosphodiesterase: a study of the EAL domain-containing RocR from Pseudomonas aeruginosa (2008), FEMS Microbiol. Lett., 282, 65-72.

Cloned(Commentary)

Cloned (Comment)	Organism
overexpression of wild-type enzymes, and mutant LipA and LipH mutant 1H8 in Escherichia coli strain BL21(DE3) in inclusion bodies, subcloning in Escherichia coli strain DH5alpha, expression of mutant enzymes in Pseudomonase aeruginosa strains PABS1 and PABST7.1	Pseudomonas aeruginosa

Protein Variants

Protein Variants	Comment	Organism
additional information	construction of a truncated lipH gene in which a 5'-deletion of 60 nucleotides encoding a 20 amino acid membrane anchor domain is replaced by a His10-tag and cloned into pET19b, resulting in plasmid pEHTHis19	Pseudomonas aeruginosa
S53P/L162G	optimizing enantioselectivity of LipA by comprehensive directed evolution approach	Pseudomonas aeruginosa
T180I/T234S	identification of a mutant LipH 1H8 with increased (S)-enantioselectivity for model substrate 2-methyldecanoic acid 4-nitrophenyl ester compared to the wild-type enzyme, overview. The mutant shows low secretion efficiency, identification of two amino acid substitutions located on the protein surface, which significantly impair lipase secretion, the amino acid substitutions T180I and T234S of lipase variant 1H8 are located in close vicinity to these cysteines, suggesting that these substitutions may impair its ability to form disulfide bonds	Pseudomonas aeruginosa

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
extracellular	the enzyme is secreted, the formation of one disulfide bond between residues C183 and C235 is essential for secretion of lipase, and residues D20 and T180 also affect secretion of lipase, overview	Pseudomonas aeruginosa	-	-

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas aeruginosa	-	genes lipA and lipH	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant wild-type LipA and LipH mutant 1H8 from Escherichia coli strain BL21(DE3) solubilized inclusion bodies by	Pseudomonas aeruginosa

Renatured (Commentary)

Renatured (Comment)	Organism
recombinant wild-type LipA and LipH mutant 1H8 from Escherichia coli strain BL21(DE3) inclusion bodies, denaturation by treatment for 1 h at 37°C with 100 mM Tris-HCl, pH 8.0, 8 M urea, followed by in vitro refolding to enzymatic activity performed for 3 h by adding equimolar ratios of LipH and sudden 10fold dilution in a buffer containing 50 mM Tris-HCl, pH 8.0, 3.5 mM CaCl2, 0.7 mM laurylmaltoside and 45% v/v glycerol	Pseudomonas aeruginosa

Renatured (Comment)

Organism

recombinant wild-type LipA and LipH mutant 1H8 from Escherichia coli strain BL21(DE3) inclusion bodies, denaturation by treatment for 1 h at 37°C with 100 mM Tris-HCl, pH 8.0, 8 M urea, followed by in vitro refolding to enzymatic activity performed for 3 h by adding equimolar ratios of LipH and sudden 10fold dilution in a buffer containing 50 mM Tris-HCl, pH 8.0, 3.5 mM CaCl2, 0.7 mM laurylmaltoside and 45% v/v glycerol

Pseudomonas aeruginosa

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
2-methyldecanoic acid 4-nitrophenyl ester + H2O	model substrate, enantioselectivity in the asymmetric hydrolysis with preference for the S-enantiomer	Pseudomonas aeruginosa	(S)-2-methyldecanoate + 4-nitrophenol + (R)-2-methyldecanoic acid 4-nitrophenyl ester	-	?

Synonyms

Synonyms	Comment	Organism
LipA	-	Pseudomonas aeruginosa
lipase	-	Pseudomonas aeruginosa
LipH	-	Pseudomonas aeruginosa