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Aqueous two‐phase systems containing urea: Influence of protein structure on protein partitioning
Author(s) -
Rämsch Christian,
Kleinelanghorst Lutz B.,
Knieps Esther A.,
Thömmes Jörg,
Kula MariaRegina
Publication year - 2000
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/(sici)1097-0290(20000705)69:1<83::aid-bit10>3.0.co;2-0
Subject(s) - chemistry , aqueous solution , lysozyme , urea , tryptophan , phase (matter) , peg ratio , protein structure , aqueous two phase system , chromatography , biochemistry , organic chemistry , amino acid , finance , economics
During recombinant E. coli fermentation with high‐expression levels inclusion bodies are often formed. Aqueous two‐phase systems have been successfully used in the presence of urea for the initial recovery step of inclusion bodies from E. coli . Basic studies of the complex interactions are lacking. For a systematic study of protein partitioning in the presence of urea we selected T4‐lysozyme mutants with different thermal stability as a model. The stabilization of these variants by phase components was investigated measuring the fluorescence emission of tryptophan residues in the protein. Protein structure was stabilized at pH 7 in the order of S0 4 2− >> PEG = Dextran > H 2 O. The conformation of proteins was shown to have a strong influence on the partitioning in aqueous two‐phase systems. Tryptophan and its homologuous di‐ and tripeptdides were partitioned in similar phase systems to normalize for contribution from hydrophobic interactions. © 2000 John Wiley & Sons, Inc. Biotechnol Bioeng 69: 83–90, 2000.