Protein pre-fractionation in detergent–polymer aqueous two-phase systems for facilitated proteomic studies of membrane proteins

Everberg, Henrik; Sivars, Ulf; Emanuelsson, Cecilia; Persson, Cecilia; Englund, Ann-Kristin; Haneskog, Lars; Lipniunas, Peter; Jörntén‐Karlsson, Magnus; Tjerneld, Folke

doi:10.1016/j.chroma.2003.12.016

Cited by 41 publications

(18 citation statements)

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“…More recently, Everberg et al [143] proposed a novel method for prefractionation of membrane proteins, via detergent-based aqueous two-phase partitioning. This two-phase system partitioning applied to the separation of proteins was pioneered by Kula [144]; it consists of two aqueous solutions, at least one of them containing a water-soluble polymer.…”

Section: Special Focus On Membrane Proteinsmentioning

confidence: 99%

Prefractionation techniques in proteome analysis: The mining tools of the third millennium

et al. 2005

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The present review deals with prefractionation protocols used in proteomic investigation in preparation for mass spectrometry (MS) or two-dimensional electrophoresis (2-DE) map analysis. Briefly, reported methods focus on cell organelle differential centrifugation and on chromatographic approaches, to continue in extenso with a panoply of electrophoretic methods. In the case of chromatography, procedures useful as a prefractionation step, including affinity, ion-exchange, and reversed-phase resins, revealed several hundreds of new species, previously undetected in unfractionated samples. Novel chromatographic prefractionation methods are also discussed such as a multistaged fractionation column, consisting in a set of immobilized chemistries, serially connected in a stack format (an assembly of seven blocks), each capable of harvesting a given protein population. Such a method significantly simplifies the complexity of treated samples while concentrating species, all resulting in a larger number of visible proteins by MS or 2-DE. Electrophoretic prefractionation protocols include all those electrokinetic methodologies which are performed in free solution, essentially all relying on isoelectric focusing steps (although some approaches based on gels and granulated media are also discussed). Devices associated with electrophoretic separation are multichamber apparatus, such as the multicompartment electrolyzers equipped with either isoelectric membranes or with isoelectric beads. Multicup device electrophoresis and several others, exploiting the conventional technique of carrier ampholyte focusing, are reviewed. This review also reports approaches for sample treatments in order to detect low-abundance species. Among others, a special emphasis is made on the reduction of concentration difference between proteins constituting a sample. This latter consists in a library of combinatorial ligands coupled to small beads. Such a library comprises hexameric ligands composed of 20 amino acids, resulting in millions of different structures. When these beads are impregnated with complex proteomes (e.g., human sera) of widely differing protein compositions, they are able to significantly reduce the concentration differences, thus greatly enhancing the possibility to evidence low-abundance species. It is felt that this panoply of methods could offer a strong step forward in "mining below the tip of the iceberg" for detecting the "unseen proteome".

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Section: Special Focus On Membrane Proteinsmentioning

confidence: 99%

Prefractionation techniques in proteome analysis: The mining tools of the third millennium

et al. 2005

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“…In comparison with sequence tags and genomic databases, the presence of fully novel or multiple, closely related proteins can be demonstrated. Studies to test this approach have been carried out on several types of membranes, such as mitochondrial membranes from the yeast Saccharomyces cerevisiae (Everberg et al, 2004). In this paper, we will focus on photosynthetic membrane proteins from oxygen evolving organisms.…”

Section: Introductionmentioning

confidence: 99%

Single particle electron microscopy in combination with mass spectrometry to investigate novel complexes of membrane proteins

Arteni

Nowaczyk

Lax

et al. 2005

Journal of Structural Biology

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Single particle electron microscopy in combination with mass spectrometry to investigate novel complexes of membrane proteins Arteni, Ana A.; Nowaczyk, Marc; Lax, Julia; Rögner, Matthias; Boekema, Egbert J.; Kouril, R.; Rogner, M. Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.Downloaded from the University of Groningen/UMCG research database (Pure): http://www.rug.nl/research/portal. For technical reasons the number of authors shown on this cover page is limited to 10 maximum. AbstractLarge data sets of molecular projections of the membrane proteins Photosystem I and Photosystem II from cyanobacteria were analyzed by single particle electron microscopy (EM). Analysis resulted in the averaging of 2D projections from the puriWed complexes but also in the simultaneous detection and averaging of 2D projections from large contaminating complexes, which were present in frequencies as low as 0.1%. Among them T-shaped and L-shaped contaminants were found. The L-shaped particles could be assigned to Complex I just from the shape, although no Complex I from a cyanobacterium has been structurally characterized. A systematic comparison by single particle EM and mass spectrometry of two diVerently puriWed Photosystem II complexes resulted in the assignment of PsbZ, a small peripheral subunit of 6.8 kDa, within the structure. Together these data suggest that screening for membrane protein structures by single particle EM and mass spectrometry may be a new approach to Wnd novel structures of such proteins. We propose here a scheme for searching for novel membrane protein structures in speciWc types of membranes. In this approach single particle EM and mass spectrometry, after pre-fractionation using one-or multidimensional protein separation techniques, are applied to characterize all its larger components. 

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“…Everberg et al [43] isolated mitochondria proteins from the yeast S. cerevisiae using a detergent -polymer aqueous two-phase system that consisted of PEG and either one of the two commonly used nonionic detergents, Triton X-114 and dodecyl maltoside. Most recently, a two-phase on-membrane digestion method (mixed n-butanol and 25 mM NH 4 HCO 3 ) was developed by Zhou et al [44] for the preconcentration of rat liver membrane proteome, followed by capillary LC-MS/MS (CapLC-MS/MS) analysis.…”

Section: Preconcentration Of Membrane Proteinsmentioning

confidence: 99%

Microseparation of membrane proteins

Zhang¹,

Feng²,

Du³

et al. 2009

J of Separation Science

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Membrane proteins are generally of natural low abundance and insoluble to aqueous solutions. Thus, investigation of membrane proteins is relatively hampered since efficient separation of membrane proteins are rather challenging. Microseparation approaches certainly play an essential role in fulfilling such demanding investigations due to their significant advantages of high throughput, reduced separation time, and low sample consumption. In this review, recent developments of microseparation are documented with aspects of three key separation methods, including CE, micro-LC and microchip. Preparation of membrane proteins prior to separation is also discussed, including solubilization and preconcentration. Certainly, more applications of microseparation approaches in membrane protein separations can be expected in the near future.

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Protein pre-fractionation in detergent–polymer aqueous two-phase systems for facilitated proteomic studies of membrane proteins

Cited by 41 publications

References 50 publications

Prefractionation techniques in proteome analysis: The mining tools of the third millennium

Prefractionation techniques in proteome analysis: The mining tools of the third millennium

Single particle electron microscopy in combination with mass spectrometry to investigate novel complexes of membrane proteins

Microseparation of membrane proteins

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