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Investigating pH and Cu (II) effects on lipase activity and enantioselectivity via kinetic and spectroscopic methods
- Source :
- Biotechnology journal. 1(11)
- Publication Year :
- 2006
-
Abstract
- For Candida rugosa lipase (CRL) catalyzed hydrolysis of racemic 1-phenethyl acetate, both the weakly acidic pH (pH 6.0) and the addition of 1 mM copper (II) ion enhanced the enzyme activity and enantioselectivity (E value) about twofold, as compared with that under neutral pH and noadditive conditions. The decrease of activation free energy (DeltaG) and increase of k(cat)(R)/k(cat)(S) at weakly acidic pH and/or in the presence of copper (II) characterized the kinetic behavior of CRL. On the other hand, for providing reasonable insights into the catalytic mechanism and the structural basis for enantioselectivity alteration, spectroscopic techniques were employed to probe conformational changes of the enzyme in each medium assayed. The fluorescence emission spectra revealed that pH and copper (II) might exert different effects on the microenvironment of Trp residue and thereby on the protein conformation, which could be further verified by UV-visible and Raman spectra. The conformational modulation of CRL associated with either pH or copper (II) concentration in the reaction medium could be attributed to the flexible and sensitive conformation of the enzyme, which is responsible for the significant variation of apparent activity and enantioselectivity with the tuning of biocatalyst microenvironment.
- Subjects :
- Stereochemistry
Ultraviolet Rays
chemistry.chemical_element
Acetates
Spectrum Analysis, Raman
Applied Microbiology and Biotechnology
Catalysis
Hydrolysis
Protein structure
Lipase
Candida
chemistry.chemical_classification
biology
Stereoisomerism
General Medicine
Hydrogen-Ion Concentration
Copper
Enzyme assay
Kinetics
Enzyme
Spectrometry, Fluorescence
chemistry
Models, Chemical
Biocatalysis
Spectrophotometry
biology.protein
Molecular Medicine
Thermodynamics
Electrophoresis, Polyacrylamide Gel
Subjects
Details
- ISSN :
- 18607314
- Volume :
- 1
- Issue :
- 11
- Database :
- OpenAIRE
- Journal :
- Biotechnology journal
- Accession number :
- edsair.doi.dedup.....57ac51120d4549e543f335ee4ac2e8d0