Details
| Original language | English |
|---|---|
| Pages (from-to) | 150-159 |
| Number of pages | 10 |
| Journal | Enzyme and microbial technology |
| Volume | 29 |
| Issue number | 2-3 |
| Publication status | Published - 17 Jul 2001 |
Abstract
With the use of two fluorescence spectroscopic detectable substrates, L-phenylalanin-7-amido-4-methylcoumarin (L-PheAMC) and D-phenylalanin-7-amido-4-trifluoromethylcoumarin (D-PheAFC), it was possible to monitor a quasi-enantiomeric enzymatic reaction. The simultaneous hydrolysis of L-PheAMC and D-PheAFC was applied in aqueous media and via on-line 2D-fluorescence spectroscopy it was possible to draw conclusions about the enantioselectivity of the used enzymes α-chymotrypsin and esterase from porcine liver. The kinetic parameters from the single-hydrolysis of L-PheAMC and D-PheAFC were determined via a timescan run, the results showed, that the L-substrate was hydrolyzed 106 times faster than the D-substrate by α-chymotrypsin. In a simultaneous usage of both coumarin substrates with α-chymotrypsin, the L-coumarin was favored by the enzyme, even when the L-substrate was used in a 10 times lower concentration than the D-substrate. With the unspecific esterase, the L-substrate was first hydrolyzed again when equal concentrations of both coumarins were used. But the reaction was influenced by the presence of the D-substrate. With a 10 times lower L-substrate concentration, the D-substrate was hydrolyzed by the esterase, but the reaction was again influenced by the presence of the enantiomeric partner.
Keywords
- 2D-fluorescence, Chymotrypsin, Coumarin, Esterase, On-line-monitoring
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
- Chemical Engineering(all)
- Bioengineering
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
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In: Enzyme and microbial technology, Vol. 29, No. 2-3, 17.07.2001, p. 150-159.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - On-line monitoring of a quasi-enantiomeric reaction with two coumarin substrates via 2D-fluorescence spectroscopy
AU - Knüttel, Torsten
AU - Hartmann, Thorsten
AU - Meyer, Hartmut
AU - Scheper, Thomas
N1 - Funding information: The authors thank the Deutsche Forschungsgemeinschaft (DFG, as part of the Graduiertenkolleg) for financial support.
PY - 2001/7/17
Y1 - 2001/7/17
N2 - With the use of two fluorescence spectroscopic detectable substrates, L-phenylalanin-7-amido-4-methylcoumarin (L-PheAMC) and D-phenylalanin-7-amido-4-trifluoromethylcoumarin (D-PheAFC), it was possible to monitor a quasi-enantiomeric enzymatic reaction. The simultaneous hydrolysis of L-PheAMC and D-PheAFC was applied in aqueous media and via on-line 2D-fluorescence spectroscopy it was possible to draw conclusions about the enantioselectivity of the used enzymes α-chymotrypsin and esterase from porcine liver. The kinetic parameters from the single-hydrolysis of L-PheAMC and D-PheAFC were determined via a timescan run, the results showed, that the L-substrate was hydrolyzed 106 times faster than the D-substrate by α-chymotrypsin. In a simultaneous usage of both coumarin substrates with α-chymotrypsin, the L-coumarin was favored by the enzyme, even when the L-substrate was used in a 10 times lower concentration than the D-substrate. With the unspecific esterase, the L-substrate was first hydrolyzed again when equal concentrations of both coumarins were used. But the reaction was influenced by the presence of the D-substrate. With a 10 times lower L-substrate concentration, the D-substrate was hydrolyzed by the esterase, but the reaction was again influenced by the presence of the enantiomeric partner.
AB - With the use of two fluorescence spectroscopic detectable substrates, L-phenylalanin-7-amido-4-methylcoumarin (L-PheAMC) and D-phenylalanin-7-amido-4-trifluoromethylcoumarin (D-PheAFC), it was possible to monitor a quasi-enantiomeric enzymatic reaction. The simultaneous hydrolysis of L-PheAMC and D-PheAFC was applied in aqueous media and via on-line 2D-fluorescence spectroscopy it was possible to draw conclusions about the enantioselectivity of the used enzymes α-chymotrypsin and esterase from porcine liver. The kinetic parameters from the single-hydrolysis of L-PheAMC and D-PheAFC were determined via a timescan run, the results showed, that the L-substrate was hydrolyzed 106 times faster than the D-substrate by α-chymotrypsin. In a simultaneous usage of both coumarin substrates with α-chymotrypsin, the L-coumarin was favored by the enzyme, even when the L-substrate was used in a 10 times lower concentration than the D-substrate. With the unspecific esterase, the L-substrate was first hydrolyzed again when equal concentrations of both coumarins were used. But the reaction was influenced by the presence of the D-substrate. With a 10 times lower L-substrate concentration, the D-substrate was hydrolyzed by the esterase, but the reaction was again influenced by the presence of the enantiomeric partner.
KW - 2D-fluorescence
KW - Chymotrypsin
KW - Coumarin
KW - Esterase
KW - On-line-monitoring
UR - http://www.scopus.com/inward/record.url?scp=0035822718&partnerID=8YFLogxK
U2 - 10.1016/S0141-0229(01)00369-6
DO - 10.1016/S0141-0229(01)00369-6
M3 - Article
AN - SCOPUS:0035822718
VL - 29
SP - 150
EP - 159
JO - Enzyme and microbial technology
JF - Enzyme and microbial technology
SN - 0141-0229
IS - 2-3
ER -