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Literature summary extracted from

  • Gervais, D.; Foote, N.
    Recombinant deamidated mutants of Erwinia chrysanthemi L-asparaginase have similar or increased activity compared to wild-type enzyme (2014), Mol. Biotechnol., 56, 865-877.
    View publication on PubMed

Application

EC Number Application Comment Organism
3.5.1.1 medicine the enzyme is an important biopharmaceutical product used in the treatment of acute lymphoblastic leukaemia Dickeya chrysanthemi

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.1.1 mutants containing Asn to Asp changes Dickeya chrysanthemi

Protein Variants

EC Number Protein Variants Comment Organism
3.5.1.1 N281D mutant has approximately the same specific activity as the wild-type enzyme. Mutation results in a lower glutaminase activity compared with wild-type and the N41D mutant. Mutation imparts less stability to the enzyme at elevated temperatures. The N281D mutation causes a decrease in substrate affinity as well as a decrease in the stability profile. The deamidation at the N281 site should not be a concern during processing, storage or clinical use. The deamidated variant is active and stable under normal storage conditions Dickeya chrysanthemi
3.5.1.1 N41D mutant has approximately the same specific activity as the wild-type enzyme. Mutation conferrs a slight increase in kcat. Charge differences to the wild-type enzyme, at -1 per monomer or -4 per tetramer. The deamidation at the N41 site should not be a concern during processing, storage or clinical use. These deamidated variant is active and stable under normal storage conditions Dickeya chrysanthemi
3.5.1.1 N41D/N281D mutant enzyme has increased specific activity. Charge differences to the wild-type enzyme, at -1 per monomer or -4 per tetramer. The N281D mutation causes a decrease in substrate affinity as well as a decrease in the stability profile. The deamidation at the N41 and N281 sites should not be a concern during processing, storage or clinical use. These deamidated variant is active and stable under normal storage conditions Dickeya chrysanthemi

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.1.1 0.0291
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D Dickeya chrysanthemi
3.5.1.1 0.0291
-
 L-asparagine pH 8.6, 37°C, wild-type enzyme Dickeya chrysanthemi
3.5.1.1 0.0562
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D/N281D Dickeya chrysanthemi
3.5.1.1 0.0597
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N281D Dickeya chrysanthemi

Organic Solvent Stability

EC Number Organic Solvent Comment Organism
3.5.1.1 urea the N281D mutation results in an inability to refold after exposure to urea concentrations greater than 1.5 M, while the external N41D mutation has no impact on the stability of the enzyme Dickeya chrysanthemi

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.1 Dickeya chrysanthemi
-
-
-
3.5.1.1 Dickeya chrysanthemi NCPPB 1066
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.1.1 mutants containing Asn to Asp changes Dickeya chrysanthemi

Source Tissue

EC Number Source Tissue Comment Organism Textmining

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.1 L-asparagine + H2O
-
 Dickeya chrysanthemi L-aspartate + NH3
-
 ?
3.5.1.1 L-asparagine + H2O
-
 Dickeya chrysanthemi NCPPB 1066 L-aspartate + NH3
-
 ?

Subunits

EC Number Subunits Comment Organism
3.5.1.1 homotetramer
-
 Dickeya chrysanthemi

Synonyms

EC Number Synonyms Comment Organism
3.5.1.1 ErA
-
 Dickeya chrysanthemi

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.1.1 37
-
 assay at Dickeya chrysanthemi

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.5.1.1 49
-
 3 min, 50% loss of activity of mutant enzyme N281D and double mutant enzyme N41D/N281D, no loss of activity of wild-type enzyme and mutant enzyme N41D Dickeya chrysanthemi

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.1.1 565
-
 L-asparagine pH 8.6, 37°C, wild-type enzyme Dickeya chrysanthemi
3.5.1.1 573
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N281D Dickeya chrysanthemi
3.5.1.1 657
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D Dickeya chrysanthemi
3.5.1.1 798
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D/N281D Dickeya chrysanthemi

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.1.1 8.6
-
 assay at Dickeya chrysanthemi

pI Value

EC Number Organism Comment pI Value Maximum pI Value
3.5.1.1 Dickeya chrysanthemi wild-type and N281D enzymes have roughly the same pI, as do the N41D and the double mutant
-
 additional information

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.5.1.1 9597
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N281D Dickeya chrysanthemi
3.5.1.1 14200
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D/N281D Dickeya chrysanthemi
3.5.1.1 19420
-
 L-asparagine pH 8.6, 37°C, wild-type enzyme Dickeya chrysanthemi
3.5.1.1 22580
-
 L-asparagine pH 8.6, 37°C, mutant enzyme N41D Dickeya chrysanthemi