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Literature summary extracted from

  • Otten, L.G.; Sio, C.F.; Vrielink, J.; Cool, R.H.; Quax, W.J.
    Altering the substrate specificity of cephalosporin acylase by directed evolution of the beta-subunit (2002), J. Biol. Chem., 277, 42121-42127.
    View publication on PubMed

Protein Variants

EC Number Protein Variants Comment Organism
3.5.1.93 F229L turnover number for adipyl-7-aminodesacetoxycephalosporanic acid is nealy identical to wild-type value, 1.8fold decrease in KM-value for adipyl-7-aminodesacetoxycephalosporanic acid, 1.4fold decrease in turnover number for glutaryl-7-aminocephalosporanic acid, 1.45fold increase in Km-value for glutaryl-7-aminocephalosporanic acid as compared to wild-type enzyme Pseudomonas sp.
3.5.1.93 F375L 1.5fold increase in turnover number for adipyl-7-aminodesacetoxycephalosporanic acid, 1.2fold decrease in KM-value for adipyl-7-aminodesacetoxycephalosporanic acid, 3.9fold decrease in turnover number for glutaryl-7-aminocephalosporanic acid, 8.2fold increase in Km-value for glutaryl-7-aminocephalosporanic acid as compared to wild-type enzyme. Improved activity ratio for adipyl-7-aminodesacetoxycephalosporanic acid to glutaryl-7-aminocephalosporanic acid of the mutant enzyme is a consequence of a decreased catalytic efficiency towards glutaryl-7-aminocephalosporanic acid Pseudomonas sp.
3.5.1.93 M271V/Q291K/T374S 1.3fold increase in turnover number for adipyl-7-aminodesacetoxycephalosporanic acid, 1.4fold decrease in KM-value for adipyl-7-aminodesacetoxycephalosporanic acid, 1.2fold decrease in turnover number for glutaryl-7-aminocephalosporanic acid, 1.3fold increase in Km-value for glutaryl-7-aminocephalosporanic acid as compared to wild-type enzyme Pseudomonas sp.
3.5.1.93 N266H 1.2fold increase in turnover number for adipyl-7-aminodesacetoxycephalosporanic acid, 6.9fold decrease in KM-value for adipyl-7-aminodesacetoxycephalosporanic acid, 1.3fold decrease in turnover number for glutaryl-7-aminocephalosporanic acid, 2fold decrease in Km-value for glutaryl-7-aminocephalosporanic acid as compared to wild-type enzyme. Nearly 10fold improved catalytic efficiency on adipyl-7-aminodesacetoxycephalosporanic acid, resulting from a 50% increase in turnover-number and a 6fold decrease in KM-value without decreasing the catalytic efficiency on glutaryl-7-aminocephalosporanic acid Pseudomonas sp.
3.5.1.93 N266S 1.4fold decrease in turnover number for adipyl-7-aminodesacetoxycephalosporanic acid, 2.3fold decrease in KM-value for adipyl-7-aminodesacetoxycephalosporanic acid, 1.95fold decrease in turnover number for glutaryl-7-aminocephalosporanic acid, 2.8fold increase in Km-value for glutaryl-7-aminocephalosporanic acid as compared to wild-type enzyme Pseudomonas sp.

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.1.93 0.016
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme N266H Pseudomonas sp.
3.5.1.93 0.033
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, wild-type enzyme Pseudomonas sp.
3.5.1.93 0.043
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme M271V/Q291K/T374S Pseudomonas sp.
3.5.1.93 0.048
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme F229L Pseudomonas sp.
3.5.1.93 0.094
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme N266S Pseudomonas sp.
3.5.1.93 0.14
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme N266H Pseudomonas sp.
3.5.1.93 0.27
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme F375L Pseudomonas sp.
3.5.1.93 0.42
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme N266S Pseudomonas sp.
3.5.1.93 0.52
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme F229L Pseudomonas sp.
3.5.1.93 0.7
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme M271V/Q291K/T374S Pseudomonas sp.
3.5.1.93 0.82
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme F375L Pseudomonas sp.
3.5.1.93 0.96
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, wild-type enzyme Pseudomonas sp.

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.93 Pseudomonas sp. P07662 SY-77
-
3.5.1.93 Pseudomonas sp. SY-77 P07662 SY-77
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.93 adipyl-7-aminodesacetoxycephalosporanic acid + H2O
-
 Pseudomonas sp. deacetoxycephalosporanic acid + adipate
-
 ?
3.5.1.93 adipyl-7-aminodesacetoxycephalosporanic acid + H2O
-
 Pseudomonas sp. SY-77 deacetoxycephalosporanic acid + adipate
-
 ?
3.5.1.93 glutaryl-7-aminocephalosporanic acid + H2O
-
 Pseudomonas sp. 7-aminocephalosporanate + glutarate
-
 ?
3.5.1.93 glutaryl-7-aminocephalosporanic acid + H2O
-
 Pseudomonas sp. SY-77 7-aminocephalosporanate + glutarate
-
 ?

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.1.93 0.34
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme N266S Pseudomonas sp.
3.5.1.93 0.46
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, wild-type enzyme Pseudomonas sp.
3.5.1.93 0.47
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme F229L Pseudomonas sp.
3.5.1.93 0.55
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme N266H Pseudomonas sp.
3.5.1.93 0.61
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme M271V/Q291K/T374S Pseudomonas sp.
3.5.1.93 0.67
-
 adipyl-7-aminodesacetoxycephalosporanic acid pH 7.5, 37°C, mutant enzyme F375L Pseudomonas sp.
3.5.1.93 1.1
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme F375L Pseudomonas sp.
3.5.1.93 2.2
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme N266S Pseudomonas sp.
3.5.1.93 3.1
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme F229L Pseudomonas sp.
3.5.1.93 3.3
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme N266H Pseudomonas sp.
3.5.1.93 3.5
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, mutant enzyme M271V/Q291K/T374S Pseudomonas sp.
3.5.1.93 4.3
-
 glutaryl-7-aminocephalosporanic acid pH 7.5, 37°C, wild-type enzyme Pseudomonas sp.