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Reactivity of 3-HBA-6-hydroxylase with diethylpyrocarbonate and N-bromosuccinimide: Effect of chemical modifications on kinetic and spectral properties of the enzyme

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Title Reactivity of 3-HBA-6-hydroxylase with diethylpyrocarbonate and N-bromosuccinimide: Effect of chemical modifications on kinetic and spectral properties of the enzyme
 
Creator SUMATHI, S
DASGUPTA, D
 
Subject para-hydroxybenzoate hydroxylase
pseudomonas-fluorescens
3-hydroxybenzoate 6-hydroxylase
tertiary structure
sequence
involvement
assignment
substrate
residues
protein
 
Description The rapid inactivation of 3-HBA-6-hydroxylase by 100 mu M diethylpyrocarbonate or 40 mu M N-bromosuccinimide and protection offered by the substrate, 3-hydroxybenzoate, against these chemical modifications implicate the involvement of histidine and tryptophan in the catalytic activity of the enzyme. Inactivation of the enzyme by diethylpyrocarbonate followed pseudo-first-order kinetics, and an "n" value of 1.3 was obtained. Inactivation of the enzyme by N-bromosuccinimide was instantaneous and failed to follow pseudo-first-order kinetics. Distinct and incremental changes in the UV absorption, emission fluorescence, and near UV-CD spectra of the enzyme upon its titration with increasing concentrations of diethylpyrocarbonate or N-bromosuccinimide may be ascribed to modification and/or changes in the microenvironment of aromatic amino acid residue(s) such as tryptophan in the enzyme.
 
Publisher AMER CHEMICAL SOC
 
Date 2011-07-14T17:54:17Z
2011-12-26T12:49:06Z
2011-12-27T05:34:49Z
2011-07-14T17:54:17Z
2011-12-26T12:49:06Z
2011-12-27T05:34:49Z
2000
 
Type Article
 
Identifier BIOTECHNOLOGY PROGRESS, 16(4), 577-582
8756-7938
http://dx.doi.org/10.1021/bp000048g
http://dspace.library.iitb.ac.in/xmlui/handle/10054/4029
http://hdl.handle.net/10054/4029
 
Language en