Large-scale separation and production of engineered proteins, designed for facilitated recovery in detergent-based aqueous two-phase extraction systems

Selber, Klaus; Tjerneld, Folke; Collén, Anna; Hyytiä, Teppo; Nakari-Setälä, Tiina; Bailey, Michael; Fagerstrom, Richard M.; Kan, John; Laan, Joop van der; Penttilä, Merja; Kula, M.‐R.

doi:10.1016/s0032-9592(03)00198-5

Cited by 73 publications

(48 citation statements)

References 17 publications

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“…Hydrophobin fusion technology was originally developed for purifying proteins from fungal culture supernatants having a simple composition and a high concentration of secreted proteins (Linder et al, 2001(Linder et al, , 2002Selber et al, 2004). More recently, the utility of this ATPS technology was demonstrated for recovering hydrophobin fusions from insect cell extracts (Lahtinen et al, 2008).…”

Section: Discussionmentioning

confidence: 99%

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Hydrophobin Fusions for High-Level Transient Protein Expression and Purification inNicotiana benthamiana

et al. 2009

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Insufficient accumulation levels of recombinant proteins in plants and the lack of efficient purification methods for recovering these valuable proteins have hindered the development of plant biotechnology applications. Hydrophobins are small and surface-active proteins derived from filamentous fungi that can be easily purified by a surfactant-based aqueous two-phase system. In this study, the hydrophobin HFBI sequence from Trichoderma reesei was fused to green fluorescent protein (GFP) and transiently expressed in Nicotiana benthamiana plants by Agrobacterium tumefaciens infiltration. The HFBI fusion significantly enhanced the accumulation of GFP, with the concentration of the fusion protein reaching 51% of total soluble protein, while also delaying necrosis of the infiltrated leaves. Furthermore, the endoplasmic reticulum-targeted GFP-HFBI fusion induced the formation of large novel protein bodies. A simple and scalable surfactant-based aqueous two-phase system was optimized to recover the HFBI fusion proteins from leaf extracts. The single-step phase separation was able to selectively recover up to 91% of the GFP-HFBI up to concentrations of 10 mg mL 21 . HFBI fusions increased the expression levels of plant-made recombinant proteins while also providing a simple means for their subsequent purification. This hydrophobin fusion technology, when combined with the speed and posttranslational modification capabilities of plants, enhances the value of transient plant-based expression systems.

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Section: Discussionmentioning

confidence: 99%

“…ATPSs offer several benefits, since they are simple, rapid, and inexpensive while providing volume reduction, high capacity, and fast separations (Persson et al, 1999). Most importantly, the one-step ATPS purification is particularly attractive because it can be easily and effectively scaled up for industrial-scale protein purification Selber et al, 2004).…”

mentioning

confidence: 99%

Hydrophobin Fusions for High-Level Transient Protein Expression and Purification inNicotiana benthamiana

et al. 2009

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“…Aqueous two-phase micellar systems (ATPMS) have been widely and successfully used in the extraction and purification of biomolecules, such as proteins, 53 -55 viruses, 56 antibiotics, 57 as target protein extraction from unclarified microorganism culture, 60 large-scale separation of proteins, 58,59 DNA and nucleic acids, 60 optimized membrane solubilization, 61 partitioning of tagged and genetically engineered proteins 62,63 and enzymes. 64,65 In these systems, an aqueous surfactant solution, under appropriate conditions, spontaneously separates into two predominantly aqueous, yet immiscible, liquid phases, one of which has a greater concentration of micelles than the other.…”

Section: Aqueous Two-phase Micellar Systemsmentioning

confidence: 99%

Liquid–liquid extraction of biomolecules: an overview and update of the main techniques

Mazzola

Lopes

Hasmann

et al. 2007

J of Chemical Tech & Biotech

226

119

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Two-phase systems can be exploited in separation science for the extraction/purification of desired biomolecules. This article reviews recent experimental work on the use of aqueous two-phase polymer systems, two-phase aqueous micellar systems and reversed micellar systems for the extraction/purification of biomolecules. The experimental partitioning behavior of different biomolecules is reviewed and new results with nisin and lipopolysaccharides are presented. An extensive description of each system is presented, based on the biomolecules behavior and the tools employed to improve their extraction and purification.

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“…These researches can mainly be attributed to the brewery and beverage industry as well as to large scale protein expression [16,32,51,74,88]. Some specialities among these processes even govern the scale-up of insect cell culture processes for advanced glycosilated protein expression [49,55].…”

Section: Huge Scale Bioprocessesmentioning

confidence: 99%

Single-cell-bioreactors as end of miniaturization approaches in biotechnology: progresses with characterised bioreactors and a glance into the future

Meier

Huebner

2005

Bioprocess Biosyst Eng

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Incidents with single cells and their genesis have not been the major focus of science up to now. This fact is supported by the difficulties one faces when wanting to monitor and cultivate small populations of cells in a defined compartment under controlled conditions, in vitro. Several approaches of up- and down-scaling have often led to poorly understood results which might be better elucidated by understanding the cellular genesis as a function of its microenvironment. This review of the approaches of scale-up and scale-down processes illustrates technical possibilities and shows up their limitations with regard to obtainable data for the characterisation of cellular genesis and impact of the cellular microenvironment. For example, stem cell research advances underline the lack of information about the impact of the microenvironment on cellular development. Finally, a proposal of future research efforts is given on how to overcome this lack of data via a novel bioreactor setup.

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Large-scale separation and production of engineered proteins, designed for facilitated recovery in detergent-based aqueous two-phase extraction systems

Cited by 73 publications

References 17 publications

Hydrophobin Fusions for High-Level Transient Protein Expression and Purification inNicotiana benthamiana

Hydrophobin Fusions for High-Level Transient Protein Expression and Purification inNicotiana benthamiana

Liquid–liquid extraction of biomolecules: an overview and update of the main techniques

Single-cell-bioreactors as end of miniaturization approaches in biotechnology: progresses with characterised bioreactors and a glance into the future

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