Purification of recombinant protein by cold-coacervation of fusion constructs incorporating resilin-inspired polypeptides.

Abstract:

:Polypeptides containing between 4 and 32 repeats of a resilin-inspired sequence AQTPSSYGAP, derived from the mosquito Anopheles gambiae, have been used as tags on recombinant fusion proteins. These repeating polypeptides were inspired by the repeating structures that are found in resilins and sequence-related proteins from various insects. Unexpectedly, an aqueous solution of a recombinant resilin protein displays an upper critical solution temperature (cold-coacervation) when held on ice, leading to a separation into a protein rich phase, typically exceeding 200 mg/mL, and a protein-poor phase. We show that purification of recombinant proteins by cold-coacervation can be performed when engineered as a fusion partner to a resilin-inspired repeat sequence. In this study, we demonstrate the process by the recombinant expression and purification of enhanced Green fluorescent protein (EGFP) in E. coli. This facile purification system can produce high purity, concentrated protein solutions without the need for affinity chromatography or other time-consuming or expensive purification steps, and that it can be used with other bulk purification steps such as low concentration ammonium sulfate precipitation. Protein purification by cold-coacervation also minimizes the exposure of the target protein to enhanced proteolysis at higher temperature.

journal_name

Biotechnol Bioeng

authors

Lyons RE,Elvin CM,Taylor K,Lekieffre N,Ramshaw JA

doi

10.1002/bit.24565

subject

Has Abstract

pub_date

2012-12-01 00:00:00

pages

2947-54

issue

12

eissn

0006-3592

issn

1097-0290

journal_volume

109

pub_type

杂志文章
  • Plasmid maintenance in Escherichia coli recombinant cultures is dramatically, steadily, and specifically influenced by features of the encoded proteins.

    abstract::A set of eight closely related plasmid constructs carrying CI857-controlled recombinant genes has been used as a model to study plasmid stability in Escherichia coli, in the absence of antibiotic selection. Plasmid loss rates and relative interdivision times of plasmid-bearing cells and plasmid-free cells have been an...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:

    authors: Corchero JL,Villaverde A

    更新日期:1998-06-20 00:00:00

  • Comment on mathematical models which describe transcription and calculate the relationship between mRNA and protein expression ratio.

    abstract::Mathematical models to describe transcription (Arnold et al. (2001); Biotech Bioeng 72:548-561) and translation (Mehra et al. (2003); Biotech Bioeng 84:822-841) in bacteria are modified in order to improve reaction kinetics and to include the number of polymerase molecules that are active on the DNA, as well as to inc...

    journal_title:Biotechnology and bioengineering

    pub_type: 评论,信件

    doi:10.1002/bit.21065

    authors: Kremling A

    更新日期:2007-03-01 00:00:00

  • Expanded bed adsorption/desorption of proteins with Streamline Direct CST I adsorbent.

    abstract::Streamline Direct CST I is a new type of ion exchanger with multi-modal functional groups, specially designed for an expanded bed adsorption (EBA) process, which can capture directly the proteins from the high ionic strength feedstocks with a high binding capacity. In this study, an experimental study is carried out f...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.20952

    authors: Li P,Xiu G,Mata VG,Grande CA,Rodrigues AE

    更新日期:2006-08-20 00:00:00

  • Modeling fluid flow through irregular scaffolds for perfusion bioreactors.

    abstract::Direct perfusion of 3D tissue engineered constructs is known to enhance osteogenesis, which can be partly attributed to enhanced nutrient and waste transport. In addition flow mediated shear stresses are known to upregulate osteogenic differentiation and mineralization. A quantification of the hydrodynamic environment...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22277

    authors: Maes F,Van Ransbeeck P,Van Oosterwyck H,Verdonck P

    更新日期:2009-06-15 00:00:00

  • Fibrin gels engineered with pro-angiogenic growth factors promote engraftment of pancreatic islets in extrahepatic sites in mice.

    abstract::With a view toward reduction of graft loss, we explored pancreatic islet transplantation within fibrin matrices rendered pro-angiogenic by incorporation of minimal doses of vascular endothelial growth factor-A165 and platelet-derived growth factor-BB presented complexed to a fibrin-bound integrin-binding fibronectin d...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.25589

    authors: Najjar M,Manzoli V,Abreu M,Villa C,Martino MM,Molano RD,Torrente Y,Pileggi A,Inverardi L,Ricordi C,Hubbell JA,Tomei AA

    更新日期:2015-09-01 00:00:00

  • Sterilization of ginseng using a high pressure CO2 at moderate temperatures.

    abstract::The aim of this study was to determine the feasibility of using high pressure CO2 for sterilization of Ginseng powder, as an alternative method to conventional techniques such as gamma-irradiation and ethylene oxide. The Ginseng sample used in this study was originally contaminated with fungi and 5 x 10(7) bacteria/g ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22059

    authors: Dehghani F,Annabi N,Titus M,Valtchev P,Tumilar A

    更新日期:2009-02-01 00:00:00

  • Detachment of biomass from suspended nongrowing spherical biofilms in airlift reactors.

    abstract::In three-phase internal loop airlift reactors, the detachment of biomass from suspended biofilm pellets in the presence of bare carrier particles was investigated under nongrowth conditions. The detachment rate was dominated by collisions between bare carrier particles and biofilm pellets. The concentration of bare ca...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260460309

    authors: Gjaltema A,Tijhuis L,van Loosdrecht MC,Heijnen JJ

    更新日期:1995-05-05 00:00:00

  • A study of producing ethanol from cellulose using Clostridium thermocellum.

    abstract::The feasibility of producing ethanol in a continuous system from cellulose using Clostridirrrn thermocellum was investigated. This anaerobic and therniophilic bacterium was able to degrade cellulose directly into ethanol with acetic acid, hydrogen. and carbon dioxide as by-products of this fermentation. The fermentati...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260240106

    authors: Zertuche L,Zall RR

    更新日期:1982-01-01 00:00:00

  • Optimal production of glutathione by controlling the specific growth rate of yeast in fed-batch culture.

    abstract::The optimal of the specific growth rate was obtained with simple mathematical model in a yeast fed-batch cultures. The model was based on the mass balance around the fed-batch system and the relationship between the specific growth rate, mu, and the specific production rate of glutathione, rho(G). The optimal profile ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260380212

    authors: Shimizu H,Araki K,Shioya S,Suga K

    更新日期:1991-06-20 00:00:00

  • Affinity purification of plasmid DNA directly from crude bacterial cell lysates.

    abstract::We have shown previously that a sequence-specific DNA-binding protein based on the Lac repressor protein can isolate pre-purified DNA efficiently from simple buffer solution but our attempts to purify plasmids directly from crude starting materials were disappointing with impractically low DNA yields. We have optimize...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.21492

    authors: Darby RA,Forde GM,Slater NK,Hine AV

    更新日期:2007-12-01 00:00:00

  • On-line estimation of the biomass activity during animal-cell cultivations.

    abstract::A software sensor was developed to determine the volumetric biomass activity of animal cell cultivations on-line. It was based on the on-line estimation of the ATP-production rate from the oxygen uptake and the lactic-acid production rate. The sensor was verified for a batch culture of Vero cells, and a batch and a co...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/(SICI)1097-0290(19960720)51:2<206::AID-BIT

    authors: Dorresteijn RC,Numan KH,de Gooijer CD,Tramper J,Beuvery EC

    更新日期:1996-07-20 00:00:00

  • Growth and fermentation characteristics of new selected strains of Saccharomyces at high temperatures and high cell densities.

    abstract::Maintenance of high cell viability was the main characteristic of our new strains of thermotolerant Saccharomyces. Total sugar conversion to ethanol was observed for sugarcane juice fermentation at 38-40 degrees C in less than 10 h and without continuous aeration of the culture. Invertase activity differed among the s...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260370606

    authors: Laluce C,Palmieri MC,Lopes da Cruz RC

    更新日期:1991-03-15 00:00:00

  • High-throughput screening of chromatographic separations: IV. Ion-exchange.

    abstract::Ion-exchange (IEX) chromatography steps are widely applied in protein purification processes because of their high capacity, selectivity, robust operation, and well-understood principles. Optimization of IEX steps typically involves resin screening and selection of the pH and counterion concentrations of the load, was...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.21905

    authors: Kelley BD,Switzer M,Bastek P,Kramarczyk JF,Molnar K,Yu T,Coffman J

    更新日期:2008-08-01 00:00:00

  • Increased metal tolerance and bioaccumulation of zinc and cadmium in Chlamydomonas reinhardtii expressing a AtHMA4 C-terminal domain protein.

    abstract::The use of microalgal biomass for metal pollutant bioremediation might be improved by genetic engineering to modify the selectivity or capacity of metal biosorption. A plant cadmium (Cd) and zinc (Zn) transporter (AtHMA4) was used as a transgene to increase the ability of Chlamydomonas reinhardtii to tolerate 0.2 mM C...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.27476

    authors: Ibuot A,Webster RE,Williams LE,Pittman JK

    更新日期:2020-10-01 00:00:00

  • Bromoperoxidase activity in microplantlet suspension cultures of the macrophytic red alga Ochtodes secundiramea.

    abstract::Bromoperoxidase is an enzyme found in marine macroalgae that catalyzes the bromination of organic substrates. Photosynthetic microplantlet suspension cultures derived from the macrophytic red alga Ochtodes secundiramea were shown to possess bromoperoxidase. The optimum pH for O. secundiramea bromoperoxidase activity i...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.1129

    authors: Rorrer GL,Tucker MP,Cheney DP,Maliakal S

    更新日期:2001-09-05 00:00:00

  • Activation of microcarrier-attached lymphocytes in microgravity.

    abstract::A technology has been developed to achieve optimal attachment of adhesion-independent lymphocytes to microcarrier beads. The activation of T-lymphocytes by concanavalin A was tested under microgravity conditions in an experiment carried out in space during the first Spacelab Life Science Mission. Activation, measure...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260400815

    authors: Bechler B,Cogoli A,Cogoli-Greuter M,Müller O,Hunzinger E,Criswell SB

    更新日期:1992-10-20 00:00:00

  • Reasons for the apparent difference in the effects of produced and added ethanol on culture viability during rapid fermentation by Saccharomyces cerevisiae.

    abstract::By feeding ethanol at various high rates to low cell density cultures of Saccharomyces cerevisiae it was shown that the sharp fall in viability when ethanol is produced during rapid fermentations is in part a direct consequence of the high rate of change of extracellular ethanol concentration. Nevertheless, the fall i...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260350202

    authors: Dasari G,Worth MA,Connor MA,Pamment NB

    更新日期:1990-01-20 00:00:00

  • Aerobic phosphorus release linked to acetate uptake in bio-P sludge: process modeling using oxygen uptake rate.

    abstract::The main processes involved in enhanced biological phosphorus removal (EBPR) under anaerobic and subsequently aerobic conditions are widely described in the literature. Polyphosphate accumulating organisms (PAO) are the organisms responsible for this process. However, the mechanisms of PAO are not fully established ye...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10868

    authors: Guisasola A,Pijuan M,Baeza JA,Carrera J,Casas C,Lafuente J

    更新日期:2004-03-30 00:00:00

  • Cell cycle progression in Escherichia coli B/r affects transcription of certain genes: Implications for synthetic genome design.

    abstract::We propose that transcript levels for some genes are affected by the bacterial cell division cycle and this may be an important factor to consider when designing synthetic bacterial genomes. To test this hypothesis, transcript levels of 58 genes in Escherichia coli B/r A were determined at five times during the cell d...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22098

    authors: Echtenkamp PL,Wilson DB,Shuler ML

    更新日期:2009-02-15 00:00:00

  • Encapsulating chondrocytes in degrading PEG hydrogels with high modulus: engineering gel structural changes to facilitate cartilaginous tissue production.

    abstract::A major challenge when designing cell scaffolds for chondrocyte delivery in vivo is creating scaffolds with sufficient mechanical properties to restore initial function while simultaneously controlling temporal changes in the gel structure to facilitate tissue formation. To address this design challenge, degradable ph...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.20160

    authors: Bryant SJ,Bender RJ,Durand KL,Anseth KS

    更新日期:2004-06-30 00:00:00

  • High- and low-molecular-weight chitosan act as adjuvants during single-dose influenza A virus protein vaccination through distinct mechanisms.

    abstract::The investigation of new adjuvants is essential for the development of efficacious vaccines. Chitosan (CS), a derivative of chitin, has been shown to act as an adjuvant, improving vaccine-induced immune responses. However, the effect of CS molecular weight (MW) on this adjuvanticity has not been investigated, despite ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.27647

    authors: Lampe AT,Farris EJ,Brown DM,Pannier AK

    更新日期:2020-12-07 00:00:00

  • Cellulase production by trichoderma harzianum in static and mixed solid-state fermentation reactors under nonaseptic conditions.

    abstract::Cellulase production from lignocellulosic materials was studied in solid-state cultivation by both static and mixed techniques under nonaseptic conditions. The effects of fermentation conditions, such as moisture content, pH, temperature, and aeration, on cellulase production by Trichoderma harzianum using a mixture o...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260270917

    authors: Deschamps F,Giuliano C,Asther M,Huet MC,Roussos S

    更新日期:1985-09-01 00:00:00

  • Fluorescence-based sensing system for copper using genetically engineered living yeast cells.

    abstract::A whole cell-based optical sensing system for copper was developed based on Saccharomyces cerevisiae cells harboring plasmid pYEX-GFPuv. The basis of this system was the ability of the transcriptional activator protein Ace1 present in S. cerevisiae to control the expression of the reporter protein, GFPuv. When copper ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.20331

    authors: Shetty RS,Deo SK,Liu Y,Daunert S

    更新日期:2004-12-05 00:00:00

  • The design of controllers for batch bioreactors.

    abstract::The implementation of control algorithms to batch bioreactors is often complicated by variations in process dynamics that occur during the course of fermentation. Such a wide operating range often renders the performance of fixed gain proportional-integral-differential (PID) controllers unsatisfactory. In this work, d...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260320415

    authors: Cardello RJ,San KY

    更新日期:1988-08-05 00:00:00

  • Countercurrent multistage fluidized bed reactor for immobilized biocatalysts: I. Modeling and simulation.

    abstract::For the application of immobilized enzymes, fixed bed reactors are used almost exclusively. Fixed bed reactors have specific disadvantages, especially for processes with a deactivating catalyst. Therefore, we have studied a novel reactor type with continuous transport of the immobilized biocatalyst. Flow of biocatalys...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260360407

    authors: Vos HJ,Groen DJ,Potters JJ,Luyben KC

    更新日期:1990-08-05 00:00:00

  • Influence of uncertainties in pH, pMg, activity coefficients, metabolite concentrations, and other factors on the analysis of the thermodynamic feasibility of metabolic pathways.

    abstract::Thermodynamic feasibility analysis (TFA) has been used as a tool capable of providing additional constraints to the mass balance-based methods of analysis of metabolic networks (e.g., flux balance analysis). Several publications have recently appeared in which TFA of different metabolic pathways from relatively simple...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22309

    authors: Vojinović V,von Stockar U

    更新日期:2009-07-01 00:00:00

  • Understanding viral partitioning in two-phase aqueous nonionic micellar systems: 1. Role of attractive interactions between viruses and micelles.

    abstract::The partitioning behavior of viruses in the two-phase aqueous nonionic n-decyl tetra(ethylene oxide) (C10E4) micellar system cannot be fully explained by considering solely the repulsive, steric, excluded-volume interactions that operate between the viruses and the nonionic C10E4 micelles. Specifically, an excluded-vo...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10193

    authors: Kamei DT,Liu CL,Haase-Pettingell C,King JA,Wang DI,Blankschtein D

    更新日期:2002-04-20 00:00:00

  • Real-time dissolved carbon dioxide monitoring II: Surface aeration intensification for efficient CO2 removal in shake flasks and mini-bioreactors leads to superior growth and recombinant protein yields.

    abstract::Mass transfer is known to play a critical role in bioprocess performance and henceforth monitoring dissolved O2 (DO) and dissolved CO2 (dCO2 ) is of paramount importance. At bioreactor level these parameters can be monitored online and can be controlled by sparging air/oxygen or stirrer speed. However, traditional sma...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.27252

    authors: Chopda VR,Holzberg T,Ge X,Folio B,Wong L,Tolosa M,Kostov Y,Tolosa L,Rao G

    更新日期:2020-04-01 00:00:00

  • Heat and mass transfer effects in static solid-substrate fermentations: design of fermentation chambers.

    abstract::An experimental device was constructed to allow nearly simultaneous measurements to be made on temperature and gas composition at different depths in a solid-substrate fermentation bed. The time-dependent values of temperature, mol % O(2) and mol % CO(2) were measured at five positions in beds 6.35 cm (2.5 in.) deep. ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260250405

    authors: Rathbun BL,Shuler ML

    更新日期:1983-04-01 00:00:00

  • New approaches for verification of kinetic parameters of immobilized concanavalin A: invertase preparations investigated by flow microcalorimetry.

    abstract::In our preceding article, we demonstrated a procedure based upon enzymic flow microcalorimetry using an enzyme thermistor (ET) to characterize the microkinetic properties of an immobilized enzyme (IME) and its further application in the screening of IMEs. To consider the ET method (single ET unit, ET system 1) as stan...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/(SICI)1097-0290(19960105)49:1<26::AID-BIT4

    authors: Gemeiner P,Docolomansky P,Nahálka J,Stefuca V,Danielsson B

    更新日期:1996-01-05 00:00:00