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Literature summary extracted from

  • Narasimhan, D.; Nance, M.; Gao, D.; Ko, M.; MacDonald, J.; Tamburi, P.; Yoon, D.; Landry, D.; Woods, J.; Zhan, C.; Tesmer, J.; Sunahara, R.
    Structural analysis of thermostabilizing mutations of cocaine esterase (2010), Protein Eng., 23, 537-547.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
3.1.1.84 medicine cocaine esterase is used to accelerate the removal of systemic cocaine and to prevent cocaine-induced lethality Rhodococcus sp.

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.1.1.84 expressed in Escherichia coli BL-21 Gold (DE3) cells Rhodococcus sp.

Protein Variants

EC Number Protein Variants Comment Organism
3.1.1.84 G173Q the mutant does not have any deleterious effects on the catalytic efficiency Rhodococcus sp.
3.1.1.84 L169K the mutation significantly increases the stability of cocaine esterase over that of wild type enzyme (half-life at 37°C is 570 min). The mutant exhibits about 8fold increase in Km for cocaine compared to the wild type enzyme Rhodococcus sp.
3.1.1.84 T172R the mutants shows about wild type thermal stability and decreased catalytic efficiency for cocaine Rhodococcus sp.
3.1.1.84 T172R/G173Q the mutation extends half-life at 37°C up to 370 min (30fold improvement compared to the wild type stability) and leads to about 3fold decrease of catalytic efficiency for cocaine Rhodococcus sp.
3.1.1.84 T172R/G173Q/L169K the mutant shows poor enzyme kinetics and does not display enhanced stabilization Rhodococcus sp.

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.1.1.84 0.0037
-
 cocaine mutant enzyme G173Q, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 0.0057
-
 cocaine wild type enzyme, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 0.017
-
 cocaine mutant enzyme T172R, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 0.017
-
 cocaine mutant enzyme T172R/G13Q, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 0.044
-
 cocaine mutant enzyme L169K, at pH 7.4 and 37°C Rhodococcus sp.

Organism

EC Number Organism UniProt Comment Textmining
3.1.1.84 Rhodococcus sp. Q9L9D7
-
-
3.1.1.84 Rhodococcus sp. MB1 Q9L9D7
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.1.1.84 Talon metal chelate affinity column chromatography and Q-Sepharose column chromatography Rhodococcus sp.

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.1.1.84 cocaine + H2O
-
 Rhodococcus sp. ecgonine methyl ester + benzoate
-
 ?
3.1.1.84 cocaine + H2O
-
 Rhodococcus sp. MB1 ecgonine methyl ester + benzoate
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.1.1.84 cocE
-
 Rhodococcus sp.

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.1.1.84 37
-
 the wild type enzyme has a half-life of 12.2 min at 37°C Rhodococcus sp.

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.1.1.84 36.1
-
 cocaine mutant enzyme G173Q, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 50.9
-
 cocaine mutant enzyme T172R, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 51.4
-
 cocaine wild type enzyme, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 53.4
-
 cocaine mutant enzyme T172R/G13Q, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 80.1
-
 cocaine mutant enzyme L169K, at pH 7.4 and 37°C Rhodococcus sp.

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.1.1.84 1840
-
 cocaine mutant enzyme L169K, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 3060
-
 cocaine mutant enzyme T172R, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 3180
-
 cocaine mutant enzyme T172R/G13Q, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 8990
-
 cocaine wild type enzyme, at pH 7.4 and 37°C Rhodococcus sp.
3.1.1.84 9880
-
 cocaine mutant enzyme G173Q, at pH 7.4 and 37°C Rhodococcus sp.