L-Tryptophan → D-Tryptophan

HRID 658075Notes

Reaction equation

HRID 658075 reaction equation
[NH2:1][C@H:2]([C:13]([OH:15])=[O:14])[CH2:3][C:4]1[C:12]2[C:7](=[CH:8][CH:9]=[CH:10][CH:11]=2)[NH:6][CH:5]=1.P([O-])([O-])([O-])=O.[K+].[K+].[K+]>>[NH2:1][C@@H:2]([C:13]([OH:15])=[O:14])[CH2:3][C:4]1[C:12]2[C:7](=[CH:8][CH:9]=[CH:10][CH:11]=2)[NH:6][CH:5]=1

Participants

3

Reactant 2

HCID 6305L-Tryptophan2×HCID 62657Potassium Phosphate, Tribasic

Product 1

HCID 9060D-Tryptophan

Procedure

Total protein assays were done using a Pierce BCA kit (Rockford, Ill.). Tryptophan racemase assays with the mutant enzyme were performed in triplicate using the wild-type enzyme prepared in the same manner as a positive control. The assays contained per mL: 30 mM L-tryptophan, 50 mM potassium phosphate pH 8, 10 μM PLP, and approximately 200 μg of racemase protein in a cell free extract. Zero, 20 minute, 1 hour, and overnight samples were collected, treated with 2% formic acid, filtered, and diluted 1:10 for analysis using the chiral amino acid method described in Example 1. At the 20 minute time point, the Y396C mutant produced approximately 264 ppm D-tryptophan, as compared to 761 ppm for the wild-type enzyme. At 1 hour, the mutant produced an average of 444 ppm D-tryptophan, as compared to 1110 ppm for the wild-type enzyme. The overnight samples contained 2300 ppm D-tryptophan, whereas the wild-type enzyme had reached equilibrium by producing approximately 3000 ppm D-tryptophan.

Workup

  1. customprepared in the same manner as a positive control
  2. customZero, 20 minute, 1 hour, and overnight samples were collected
  3. additiontreated with 2% formic acid
  4. filtrationfiltered
  5. additiondiluted 1:10 for analysis

Sources & evidence

ORD record
ord-1c0ff213f88d4a3bb8ca69dda6eb78f6
Source dataset
uspto-grants-2011_12
Patent number
US08076108B2

Reaction notes

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