Effects of fused tryptophan rich peptides to a recombinant protein A domain on the partitioning in polyethylene glycol-dextran and Ucon-dextran aqueous two-phase systems

Mats Carlsson, Kristina Berggren, Per Linse, Andres Veide, Folke Tjerneld

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25 Citations (SciVal)

Abstract

Genetic engineering has been used to construct fusion proteins with tryptophan containing peptides. The peptides and the fusion proteins have been partitioned in aqueous two-phase systems of poly(ethylene glycol) (PEG)-dextran and Ucon-dextran. The studied model protein was ZZT0, where Z is an engineered domain of domain B of staphylococcal protein A. The specially designed hydrophobic peptides, Ala-Trp-Trp-Pro (T1) and (Ala-Trp-Trp-Pro)2 (T2), have been inserted into ZZT0, to give the peptide-protein fusions ZZT1 and ZZT2. In the experimental studies it was found that T1 and T2 preferred the PEG phase and even more the Ucon phase over the dextran phase. For T2 the partitioning was more one sided than for T1. For the fusion proteins, ZZT1 and ZZT2, the partitioning was enhanced into the PEG or Ucon rich phase as compared to ZZT0. The effects were lower than expected from independent contributions to the partition coefficient from the protein and the peptides. A heterogeneous lattice model was used to calculate theoretical peptide and protein partition coefficients. The calculations could reproduce the qualitative features of the experimental data. The model results suggest that a part of these experimentally observed effects is due to a depletion zone, i.e. a zone of reduced polymer concentration around the protein. The experimental results indicate a further reduction of the partition coefficient, beyond that predicted by the lattice calculations. A possible folding of the inserted peptide is discussed as a plausible mechanism for this further reduction in the partition coefficient.
Original languageEnglish
Pages (from-to)107-117
JournalJournal of Chromatography A
Volume756
Issue number1-2
DOIs
Publication statusPublished - 1996

Subject classification (UKÄ)

  • Biological Sciences
  • Physical Chemistry

Keywords

  • Partitioning
  • Aqueous two-phase systems
  • Thermoseparating polymer
  • Heterogeneous lattice model
  • Protein A

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