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Studies on the isolation of penicillin acylase from Escherichia coli by aqueous two‐phase partitioning
Author(s) -
Guan Yue,
Wu XingYan,
Treffry Timothy E.,
Lilley Terence H.
Publication year - 1992
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260400410
Subject(s) - peg ratio , chemistry , polyethylene glycol , aqueous solution , chromatography , penicillin amidase , trimethylamine , extraction (chemistry) , aqueous two phase system , potassium phosphate , enzyme , immobilized enzyme , biochemistry , organic chemistry , finance , economics
A method of enzyme release and aqueous two‐phase extraction is described for the separation of penicillin acylase from Escherichia coli cells. Butyl acetate, 12% (v/v), treatment combined with freeze–thawing gives up to 70% enzyme release. For polyethylene glycol (PEG) + phosphate two‐phase extraction systems the enzyme purity and yield were rather low. Modified PEG, including PEG–ampicillin, PEG–aniline, PEG–phosphate, and PEG–trimethylamine, were synthesized and used in aqueous two‐phase systems; PEG–trimethylamine is the most satisfactory. A system containing 12% (w/w) PEG4000, 8% (w/w) of which is PEG–trimethylamine, with 0.7 M potasium phosphate at pH 7.2, resulted in the enzyme selective partition being greatly enhanced by charge directed effects. Possible mechanisms for the separation process are discussed. © 1992 John Wiley & Sons, Inc.