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Using capillary electrophoresis in the optimization of a carboxypeptidase Y catalyzed transpeptidation reaction
Author(s) -
Vinther Anders,
Adelhorst Kim,
Kirk Ole
Publication year - 1993
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150140175
Subject(s) - capillary electrophoresis , chemistry , carboxypeptidase , peptide , chromatography , substrate (aquarium) , amide , catalysis , enzyme , carboxypeptidase a , electrophoresis , stereochemistry , biochemistry , biology , ecology
A peptide amide, R‐Arg‐NH 2 , was produced by carboxypeptidase Y (CPDY)‐catalyzed transpeptidation of a peptide, R‐Ala‐OH in presence of a large excess of Arg‐NH 2 . Baseline separation of R‐Ala‐OH and R‐Arg‐NH 2 was achieved by free solution capillary electrophoresis (CE) analysis. With CE the reactions could be closely followed with an analysis frequency of 3–6 h −1 . Due to a low consumption of sample per CE analysis (1–5 nL introduced, 5–6 μL in the sample vial), the reactions were performed in 100 and 250 μL volumes. Consequently, the optimization experiments consumed limited amounts of enzyme and substrate only. At optimized experimental conditions approximately 90% conversion of the starting peptide, R‐Ala‐OH, to R‐Arg‐NH 2 was achieved.