Investigating pH and Cu (II) effects on lipase activity and enantioselectivity via kinetic and spectroscopic methods

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 conditio...

Full description

Saved in:
Bibliographic Details
Published in:Biotechnology journal 2006-11, Vol.1 (11), p.1293-1301
Main Authors: Xu, Tian-Wen, Xu, Jian-He, Yu, Wei, Zhong, Ju-Hua
Format: Article
Language:English
Subjects:
1-Phenethyl acetate
Acetates - chemistry
Candida - enzymology
Candida rugosa
Candida rugosa lipase
Conformation modulation
Copper - chemistry
Electrophoresis, Polyacrylamide Gel
Enantioselectivity
Hydrogen-Ion Concentration
Hydrolysis
Hydrolytic activity
Kinetics
Lipase - chemistry
Lipase - metabolism
Models, Chemical
Spectrometry, Fluorescence
Spectrophotometry - methods
Spectrum Analysis, Raman
Stereoisomerism
Thermodynamics
Ultraviolet Rays
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary: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 (ΔG) and increase of kcat(R)/kcat(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.
ISSN:1860-6768
1860-7314
DOI:10.1002/biot.200600135