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Thermoseparating water/polymer system: A novel one-polymer aqueous two-phase system for protein purification

Johansson, Hans-Olof LU ; Persson, Josefine and Tjerneld, Folke LU (1999) In Biotechnology and Bioengineering 66(4). p.247-257
Abstract
In this study we show that proteins can be partitioned and separated in a novel aqueous two-phase system composed of only one polymer in water solution. This system represents an attractive alternative to traditional two-phase systems which uses either two polymers (e.g., PEG/dextran) or one polymer in high-salt concentration (e.g., PEG/salt). The polymer in the new system is a linear random copolymer composed of ethylene oxide and propylene oxide groups which has been hydrophobically modified with myristyl groups (C14H29) at both ends (HM-EOPO). This polymer thermoseparates in water, with a cloud point at 14degreesC. The HM-EOPO polymer forms an aqueous two-phase system with a top phase composed of almost 100% water and a bottom phase... (More)
In this study we show that proteins can be partitioned and separated in a novel aqueous two-phase system composed of only one polymer in water solution. This system represents an attractive alternative to traditional two-phase systems which uses either two polymers (e.g., PEG/dextran) or one polymer in high-salt concentration (e.g., PEG/salt). The polymer in the new system is a linear random copolymer composed of ethylene oxide and propylene oxide groups which has been hydrophobically modified with myristyl groups (C14H29) at both ends (HM-EOPO). This polymer thermoseparates in water, with a cloud point at 14degreesC. The HM-EOPO polymer forms an aqueous two-phase system with a top phase composed of almost 100% water and a bottom phase composed of 5-9% HM-EOPO in water when separated at 17-30degreesC. The copolymer is self-associating and forms micellar-like structures with a CMC at 12 µM (0.01%). The partitioning behavior of three proteins (lysozyme, bovine serum albumin, and apolipoprotein A-1) in water/HM-EOPO two-phase systems has been studied, as well as the effect of various ions, pH, and temperature on protein partitioning. The amphiphilic protein apolipoprotein A-1 was strongly partitioned to the HM-EOPO-rich phase within a broad-temperature range. The partitioning of hydrophobic proteins can be directed with addition of salt. Below the isoelectric point (pI) BSA was partitioned to the HM-EOPO-rich phase and above the pI to the water phase when NaClO4was added to the system. Lysozyme was directed to the HM-EOPO phase with NaClO4, and to the water phase with Na-phosphate. The possibility to direct protein partitioning between water and copolymer phases shows that this system can be used for protein separations. This was tested on purification of apolipoprotein A-1 from human plasma and Escherichia coli extract. Apolipoprotein A-1 could be recovered in the HM-EOPO-rich phase and the majority of contaminating proteins in the water phase. By adding a new water/buffer phase at higher pH and with 100 mM NaClO4, and raising the temperature for separation, the apolipoprotein A-1 could be back-extracted from the HM-EOPO phase into the new water phase. This novel system has a strong potential for use in biotechnical extractions as it uses only one polymer and can be operated at moderate temperatures and salt concentrations and furthermore, the copolymer can be recovered. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 66: 247-257, 1999. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
aqueous two-phase systems, thermoseparating polymers, polymer recycling, protein purification
in
Biotechnology and Bioengineering
volume
66
issue
4
pages
247 - 257
publisher
John Wiley & Sons
external identifiers
  • scopus:0033396212
ISSN
1097-0290
language
English
LU publication?
yes
id
a423bc4f-f991-4ea9-b0b7-15abf18baeb9 (old id 125341)
alternative location
http://www3.interscience.wiley.com/cgi-bin/abstract/71003618/ABSTRACT
http://www3.interscience.wiley.com/cgi-bin/fulltext/71003618/PDFSTART
date added to LUP
2007-07-05 15:56:38
date last changed
2017-06-25 03:46:32
@article{a423bc4f-f991-4ea9-b0b7-15abf18baeb9,
  abstract     = {In this study we show that proteins can be partitioned and separated in a novel aqueous two-phase system composed of only one polymer in water solution. This system represents an attractive alternative to traditional two-phase systems which uses either two polymers (e.g., PEG/dextran) or one polymer in high-salt concentration (e.g., PEG/salt). The polymer in the new system is a linear random copolymer composed of ethylene oxide and propylene oxide groups which has been hydrophobically modified with myristyl groups (C14H29) at both ends (HM-EOPO). This polymer thermoseparates in water, with a cloud point at 14degreesC. The HM-EOPO polymer forms an aqueous two-phase system with a top phase composed of almost 100% water and a bottom phase composed of 5-9% HM-EOPO in water when separated at 17-30degreesC. The copolymer is self-associating and forms micellar-like structures with a CMC at 12 µM (0.01%). The partitioning behavior of three proteins (lysozyme, bovine serum albumin, and apolipoprotein A-1) in water/HM-EOPO two-phase systems has been studied, as well as the effect of various ions, pH, and temperature on protein partitioning. The amphiphilic protein apolipoprotein A-1 was strongly partitioned to the HM-EOPO-rich phase within a broad-temperature range. The partitioning of hydrophobic proteins can be directed with addition of salt. Below the isoelectric point (pI) BSA was partitioned to the HM-EOPO-rich phase and above the pI to the water phase when NaClO4was added to the system. Lysozyme was directed to the HM-EOPO phase with NaClO4, and to the water phase with Na-phosphate. The possibility to direct protein partitioning between water and copolymer phases shows that this system can be used for protein separations. This was tested on purification of apolipoprotein A-1 from human plasma and Escherichia coli extract. Apolipoprotein A-1 could be recovered in the HM-EOPO-rich phase and the majority of contaminating proteins in the water phase. By adding a new water/buffer phase at higher pH and with 100 mM NaClO4, and raising the temperature for separation, the apolipoprotein A-1 could be back-extracted from the HM-EOPO phase into the new water phase. This novel system has a strong potential for use in biotechnical extractions as it uses only one polymer and can be operated at moderate temperatures and salt concentrations and furthermore, the copolymer can be recovered. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 66: 247-257, 1999.},
  author       = {Johansson, Hans-Olof and Persson, Josefine and Tjerneld, Folke},
  issn         = {1097-0290},
  keyword      = {aqueous two-phase systems,thermoseparating polymers,polymer recycling,protein purification},
  language     = {eng},
  number       = {4},
  pages        = {247--257},
  publisher    = {John Wiley & Sons},
  series       = {Biotechnology and Bioengineering},
  title        = {Thermoseparating water/polymer system: A novel one-polymer aqueous two-phase system for protein purification},
  volume       = {66},
  year         = {1999},
}