Spectroscopic investigation of lipase from Pseudomonas cepacia solubilized in 1,4-dioxane by non-covalent complexation with methoxypoly(ethylene glycol)

Lipase from Pseudomonas cepacia was made soluble in 1,4‐dioxane by lyophilization of the enzyme from aqueous solutions containing methoxypoly(ethylene glycol) (PEG). The solubility of the enzyme–PEG complex depended both on protein concentration and PEG protein ratio. Intrinsic protein fluorescence...

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Bibliographic Details
Published in:Biotechnology and bioengineering 1999-09, Vol.64 (5), p.624-629
Main Authors: Secundo, Francesco, Carrea, Giacomo, Vecchio, Giuseppe, Zambianchi, Francesca
Format: Article
Language:English
Subjects:
1,4-Dioxane
4-dioxane
Biological and medical sciences
Biotechnology
circular dichroism
Complexation
Enzyme engineering
Enzymes
Fluorescence
Fundamental and applied biological sciences. Psychology
lipase
Methods. Procedures. Technologies
methoxypoly(ethylene glycol)
methoxypolyethylene glycol
Microorganisms
Miscellaneous
Organic compounds
Organic solvents
Proteins
Pseudomonas cepacia
Solubility
Solutions
Spectroscopic analysis
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Summary:Lipase from Pseudomonas cepacia was made soluble in 1,4‐dioxane by lyophilization of the enzyme from aqueous solutions containing methoxypoly(ethylene glycol) (PEG). The solubility of the enzyme–PEG complex depended both on protein concentration and PEG protein ratio. Intrinsic protein fluorescence and far‐ and near‐UV circular dichroism revealed that not only did the enzyme not unfold in the organic solvent, but rather became more compact. This was seen by the slight quenching of fluorescence intensity and by the enhancement of the near‐UV circular dichroism negative signals, which are indicative of stronger interactions of tryptophanyl and/or tyrosyl residues among themselves or with other parts of the enzyme molecule. The specific activity of the lipase–PEG complex in the organic solvent was at least 2 orders of magnitude higher than that of the enzyme powder. This can be attributed both to the maintenance of native conformation and to enzyme dissolution in the reaction medium which should minimize possible limitations to enzyme–substrate interactions. © 1999 John Wiley & Sons, Inc., Biotechnol Bioeng 64: 624–629, 1999.
ISSN:0006-3592
1097-0290
DOI:10.1002/(SICI)1097-0290(19990905)64:5<624::AID-BIT14>3.0.CO;2-P