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Literature summary extracted from

  • Kino, K.; Sato, M.; Yoneyama, M.; Kirimura, K.
    Synthesis of DL-tryptophan by modified broad specificity amino acid racemase from Pseudomonas putida IFO 12996 (2007), Appl. Microbiol. Biotechnol., 73, 1299-1305.
    View publication on PubMed

Application

EC Number Application Comment Organism
5.1.1.10 biotechnology since commercially available D-Trp is chemically synthesized and expensive a mutant BAR protein might represent an effective method to synthesize D-Trp Pseudomonas putida

Cloned(Commentary)

EC Number Cloned (Comment) Organism
5.1.1.10 recombinant BAR is cloned in Escherichia coli BL-21 as His-tagged BAR Pseudomonas putida

Protein Variants

EC Number Protein Variants Comment Organism
5.1.1.10 I384M I384M mutant BAR shows the highest activity for Trp. Using I384M mutant BAR the proportion of D-Trp reaches 43% while wildtype BAR racemizes only 6% of initial L-Trp Pseudomonas putida
5.1.1.10 I384M specific activity: 0.124 (substrate Trp), 0.0744 (substrate Phe), 0.223 (substrate Lys), 0.026 (substrate Ala) Pseudomonas putida
5.1.1.10 I83L/D361V/Y396C specific activity: 0.0303 (substrate Trp), 0.0257 (substrate Phe), 0.473 (substrate Lys), 0.0121 (substrate Ala) Pseudomonas putida
5.1.1.10 L126H/Y396C specific activity: 0.0524 (substrate Trp), 0.0436 (substrate Phe), 0.477 (substrate Lys), 0.0144 (substrate Ala) Pseudomonas putida
5.1.1.10 additional information random mutagenesis on bar gene is performed to obtain mutant BAR derivates with high activity for Trp. Five positive mutant are isolated after the two-step-screening of the randomly mutated BAR Pseudomonas putida
5.1.1.10 additional information substitutions at Y396 and I384 increases the Trp specific racemization activity and the racemazation activity for overall amino acids, respectively Pseudomonas putida
5.1.1.10 Y293A/Y301S/Y396C specific activity: 0.0267 (substrate Trp), 0.0249 (substrate Phe), 0.458 (substrate Lys), 0.0141 (substrate Ala) Pseudomonas putida
5.1.1.10 Y396C specific activity: 0.0487 (substrate Trp), 0.00706 (substrate Phe), 0.03215 (substrate Lys), 0.825 (substrate Ala) Pseudomonas putida

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
5.1.1.10 45000
-
 SDS-PAGE Pseudomonas putida

Organism

EC Number Organism UniProt Comment Textmining
5.1.1.10 Pseudomonas putida
-
 IFO 12996
-

Purification (Commentary)

EC Number Purification (Comment) Organism
5.1.1.10 His6-tagged BAR is applied onto a His-Trap HP 1-ml column Pseudomonas putida

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
5.1.1.10 additional information
-
 wildtype BAR, specific activity for L-Lys 100%, L-Arg 65%, L-Ala 33%, L-Ser 20%, L-Met 14%, L-Cys 14%, L-Leu 3.3%, L-His 1.6%, L-Phe 0.29%, L-Pro 0.1.3%, L-Thr 0.069%, L-Asn 0.054%, L-Asp 0.01%, L-Trp 0.006%, L-Val 0.003% Pseudomonas putida
5.1.1.10 0.00588
-
 substrate Trp, wildtype BAR Pseudomonas putida
5.1.1.10 0.00706
-
 substrate Phe, mutant Y396C Pseudomonas putida
5.1.1.10 0.00782
-
 substrate Ala, wildtype BAR Pseudomonas putida
5.1.1.10 0.00838
-
 substrate Phe, wildtype BAR Pseudomonas putida
5.1.1.10 0.0121
-
 substrate Ala, mutant I83L, D361V, Y396C Pseudomonas putida
5.1.1.10 0.0141
-
 substrate Ala, mutant Y293A, Y301S, Y396C Pseudomonas putida
5.1.1.10 0.0144
-
 substrate Ala, mutant L126H, Y396C Pseudomonas putida
5.1.1.10 0.0249
-
 substrate Phe, mutant Y293A, Y301S, Y396C Pseudomonas putida
5.1.1.10 0.0257
-
 substrate Phe, mutant I83L, D361V, Y396C Pseudomonas putida
5.1.1.10 0.026
-
 substrate Ala, mutant I384M Pseudomonas putida
5.1.1.10 0.0267
-
 substrate Trp, mutant Y293A, Y301S, Y396C Pseudomonas putida
5.1.1.10 0.0303
-
 substrate Trp, mutant I83L, D361V, Y396C Pseudomonas putida
5.1.1.10 0.03215
-
 substrate Lys, mutant Y396C Pseudomonas putida
5.1.1.10 0.0436
-
 substrate Phe, mutant L126H, Y396C Pseudomonas putida
5.1.1.10 0.0487
-
 substrate Trp, mutant Y396C Pseudomonas putida
5.1.1.10 0.0524
-
 substrate Trp, mutant L126H, Y396C Pseudomonas putida
5.1.1.10 0.0744
-
 substrate Phe, mutant I384M Pseudomonas putida
5.1.1.10 0.124
-
 substrate Trp, mutant I384M Pseudomonas putida
5.1.1.10 0.132
-
 substrate Lys, wildtype BAR Pseudomonas putida
5.1.1.10 0.223
-
 substrate Lys, mutant I384M Pseudomonas putida
5.1.1.10 0.458
-
 substrate Lys, mutant Y293A, Y301S, Y396C Pseudomonas putida
5.1.1.10 0.473
-
 substrate Lys, mutant I83L, D361V, Y396C Pseudomonas putida
5.1.1.10 0.477
-
 substrate Lys, mutant L126H, Y396C Pseudomonas putida
5.1.1.10 0.825
-
 substrate Ala, mutant Y396C Pseudomonas putida

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
5.1.1.10 L-Trp
-
 Pseudomonas putida D-Trp
-
 r

Synonyms

EC Number Synonyms Comment Organism
5.1.1.10 broad specificity amino acid racemase
-
 Pseudomonas putida

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
5.1.1.10 37
-
 assay at Pseudomonas putida

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
5.1.1.10 8.5
-
 assay at Pseudomonas putida