当前位置： SCI文献检索 > PROTEIN EXPRESSION AND PURIFICATION期刊下所有文献 > Single-step purification of two functional human apolipoprotein E variants hyperexpressed in Escherichia coli.

Single-step purification of two functional human apolipoprotein E variants hyperexpressed in Escherichia coli.

Abstract：

:We have cloned, from total human liver RNA, the cDNA encoding apolipoprotein E3 (apoE3). Site-directed mutagenesis was used to obtain the cDNA encoding the apoE4 isoform, a major variant of this apolipoprotein in man. These two cDNAs were subcloned into the procaryotic expression vector pAHRS. A polyhistidine tag was added at the NH2-termini of the recombinant proteins (apoE3 and apoE4) to enable rapid purification. The resulting plasmids (pAHRS-apoE3 and pAHRS-apoE4) were introduced into the Escherichia coli strain BL21(DE3). Recombinant strains were grown at 37 degrees C in a Luria and Bertani medium and the addition of isopropyl beta-thiogalactoside resulted in the expression of large amounts of apoE protein (40.5 kDa), representing at least 15% of cellular proteins. The recombinant apoE isoforms were purified, under denaturating conditions, in one step by affinity chromatography on a Ni-chelated agarose column, yielding to about 20 mg of 96% pure protein per liter of culture. Compared to plasma apoE3 purified from human very low density lipoproteins, the two renatured recombinant apoE isoforms have the same secondary structure content, as revealed by circular dichroism measurement. Moreover, the recombinant apoE3 isoform shares similar properties for the association with lipids, compared to the human protein, indicating that the addition of the amino-terminal polyhistidine peptide does not influence the structure and the lipid binding properties of this recombinant apoE isoform. No differences in the secondary structure of recombinant apoE4 were detected, whereas this isoform presents specific reactivity with lipids. This simple and rapid procedure for the expression and the purification of functional recombinant apoE should therefore enable structural and physiological studies requiring large amounts of these apolipoproteins.

journal_name

Protein Expr Purif

journal_title

Protein expression and purification

authors

Pillot T,Barbier A,Visvikis A,Lozac'h K,Rosseneu M,Vandekerckhove J,Siest G

doi

10.1006/prep.1996.0061

subject

Has Abstract

pub_date

1996-06-01 00:00:00

pages

407-14

issue

4

eissn

1046-5928

issn

1096-0279

pii

S1046-5928(96)90061-3

journal_volume

7

pub_type

杂志文章

相关文献

PROTEIN EXPRESSION AND PURIFICATION文献大全
  • Purification and characterization of human dehydrodolychil diphosphate synthase (DHDDS) overexpressed in E. coli.

    abstract：:Protein asparagine (N)-linked glycosylation is a post-translational modification that occurs in the endoplasmic reticulum; it plays an important role in protein folding, oligomerization, quality control, sorting, and transport. Accordingly, disorders of glycosylation may affect practically every organ system. Dehydrod...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2017.02.001

    authors： Giladi M,Edri I,Goldenberg M,Newman H,Strulovich R,Khananshvili D,Haitin Y,Loewenstein A

    更新日期：2017-04-01 00:00:00

  • Expression and purification of anthrax toxin protective antigen from Escherichia coli.

    abstract：:Anthrax toxin consists of three separate proteins, protective antigen (PA), lethal factor (LF), and edema factor (EF). PA binds to the receptor on mammalian cells and facilitates translocation of EF or LF into the cytosol. PA is the primary component of several anthrax vaccines. In this study we expressed and purified...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.1996.0005

    authors： Sharma M,Swain PK,Chopra AP,Chaudhary VK,Singh Y

    更新日期：1996-02-01 00:00:00

  • Tabtoxin-resistant protein: overexpression, purification, and characterization.

    abstract：:One of the self-protection mechanisms in Pseudomonas syringae pv. tabaci, a pathogen of tobacco wildfire, is thought to be due to its tabtoxin-resistance gene (ttr). In this study, the ttr gene was inserted into an expression vector, pQE30, and successfully expressed in Escherichia coli M15 at high levels. The purifie...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.2001.1586

    authors： Liu J,Le Y,Ye B,Zhen Y,Zhu C,Shen J,Zhang R

    更新日期：2002-04-01 00:00:00

  • Isolation of alpha-lactalbumin, beta-lactoglobulin, and bovine serum albumin from cow's milk using gel filtration and anion-exchange chromatography including evaluation of their antigenicity.

    abstract：:The aim of this study was to introduce a simple, reproducible, and less expensive method for isolation of alpha-lactalbumin, beta-lactoglobulin, and bovine serum albumin from cow's milk while retaining their antigenicity. Whey (lactoserum) was obtained by isolating casein from defatted milk using hydrochloric acid. Gl...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/s1046-5928(03)00015-9

    authors： Neyestani TR,Djalali M,Pezeshki M

    更新日期：2003-06-01 00:00:00

  • Expression, purification and characterization of TMCO1 for structural studies.

    abstract：:Transmembrane and coiled-coil domains 1 (TMCO1) has a highly conserved amino acid sequence among species, indicating a critical role of TMCO1 in cell physiology. The deficiency of TMCO1 in humans is associated with cerebrofaciothoracic dysplasia (CFTD), glaucoma, osteogenesis and the occurrence of cancer. TMCO1 was re...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2020.105803

    authors： Zhang N,Tang M,Wen M,Cao Y,OuYang B

    更新日期：2021-03-01 00:00:00

  • Homologous expression of the feruloyl esterase B gene from Aspergillus niger and characterization of the recombinant enzyme.

    abstract：:The faeB gene encoding the feruloyl esterase B (FAEB) was isolated from Aspergillus niger BRFM131 genomic DNA. The faeB gene, with additional sequence coding for a C-terminal histidine tag, was inserted into an expression vector under the control of the gpd promoter and trpC terminator and expressed in a protease defi...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2004.05.019

    authors： Levasseur A,Benoit I,Asther M,Asther M,Record E

    更新日期：2004-09-01 00:00:00

  • Molecular cloning and functional expression of hemolysin from the sea anemone Actineria villosa.

    abstract：:The full-length cDNA that encodes the hemolytic toxin Avt-I, with 226 amino acids, from the venomous sea anemone Actineria villosa has been cloned using the oligo-capping method. The cDNA contains 681bp open reading frame and its predicted amino acid sequences revealed that Avt-I was basic polypeptides without cystein...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2004.12.003

    authors： Uechi G,Toma H,Arakawa T,Sato Y

    更新日期：2005-04-01 00:00:00

  • Development of a bacterial system for high yield expression of fully functional adrenal cytochrome b561.

    abstract：:Adrenal cytochrome b561 (cyt b561) is the prototypical member of an emerging family of proteins that are distributed widely in vertebrate, invertebrate and plant tissues. The adrenal cytochrome is an integral membrane protein with two b-type hemes and six predicted transmembrane helices. Adrenal cyt b561 is involved i...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2007.04.010

    authors： Liu W,Rogge CE,Kamensky Y,Tsai AL,Kulmacz RJ

    更新日期：2007-12-01 00:00:00

  • An optimized system for expression and purification of secreted bacterial proteins.

    abstract：:In this report, we describe an optimized system for the efficient overexpression, purification, and refolding of secreted bacterial proteins. Candidate secreted proteins were produced recombinantly in Escherichia coli as Tobacco Etch Virus protease-cleavable hexahistidine-c-myc eptiope fusion proteins. Without regard ...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2005.09.003

    authors： Geisbrecht BV,Bouyain S,Pop M

    更新日期：2006-03-01 00:00:00

  • Expression and purification of catalytically active human PHD3 in Escherichia coli.

    abstract：:Transcription factor HIF-1 is a key regulator in cellular adaptation to hypoxia. HIF prolyl hydroxylases (PHDs) control HIF-1 accumulation by hydroxylation dependent on molecular oxygen. Due to this regulation, PHDs have been pointed out as potential drug targets. We have purified catalytically active human PHD3 after...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2007.02.018

    authors： Fedulova N,Hanrieder J,Bergquist J,Emrén LO

    更新日期：2007-07-01 00:00:00

  • Efficient matrix-assisted refolding of the recombinant anti-staphylococcal truncated endolysin LysKCA and its structural and enzymatic description.

    abstract：:The recombinant truncated endolysin LysK consisting of two catalytic domains, N-terminal CHAP and amidase-2 (LysKCA) was overexpressed in E. coli in the form of inclusion bodies (IBs). These IBs were dissolved in 6 M solution of urea followed by the refolding process. The refolding efficacy of the dilution and matrix-...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2020.105683

    authors： Alaksandr Ž,Sergey G,Maksim P,Sergey K,Niyaz S,Uladzimir P,Mikhail S

    更新日期：2020-10-01 00:00:00

  • Purification of functional human Cl(-)/HCO(3)(-) exchanger, AE1, over-expressed in Saccharomyces cerevisiae.

    abstract：:There is no high-resolution structure for the membrane domain of the human erythrocyte anion exchanger, AE1 (Band 3). In this report, we have developed an expression and purification strategy for AE1 to be used in crystallization trials. Saccharomyces cerevisiae strain BJ5457 was transformed with an expression vector ...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2010.06.020

    authors： Bonar P,Casey JR

    更新日期：2010-11-01 00:00:00

  • Expression, proteolytic analysis, reconstitution, and crystallization of the tau60/tau91 subcomplex of yeast TFIIIC.

    abstract：:The transcription factor IIIC (TFIIIC) is a multisubunit DNA-binding factor required for promoter recognition and TFIIIB assembly on tRNA genes transcribed by RNA polymerase III. Yeast TFIIIC consists of six subunits, organized in the two globular subcomplexes tauA and tauB, which recognize two internal tDNA promoter ...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2005.06.013

    authors： Mylona A,Acker J,Fernández-Tornero C,Sentenac A,Müller CW

    更新日期：2006-02-01 00:00:00

  • Expression, purification and characterization of heterotrimeric forms of sTRAIL using a polycistronic expression vector.

    abstract：:Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which is capable of selectively inducing apoptosis of cancer cells, is a potential targeted drug for cancer therapy. The TRAIL protein induces apoptosis only in trimeric form. However, the recombinant soluble TRAIL (sTRAIL) trimer has low stability and a...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2015.08.004

    authors： Wang B,Wang Z,Yan J,Wang L,Wang Z,Wu J,Zhang H,Wu H,Kong W,Yu B,Yu X

    更新日期：2015-11-01 00:00:00

  • PEGylated murine Granulocyte-macrophage colony-stimulating factor: production, purification, and characterization.

    abstract：:Granulocyte-macrophage colony-stimulating factor (GM-CSF) regulates proliferation, differentiation, and function of hematopoietic progenitor cells. Aside from expansion of hematopoietic cells, GM-CSF has shown efficacy in other diseases, including Crohn's disease. While GM-CSF being clinically used in humans, the abil...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2005.08.014

    authors： Sainathan SK,Tu L,Bishnupuri KS,Han M,Li A,Newberry RD,McDonald KG,Crimmins DL,Houchen C,Anant S,Dieckgraefe BK

    更新日期：2005-12-01 00:00:00

  • Overexpression, oxidative refolding, and zinc binding of recombinant forms of the murine S100 protein MRP14 (S100A9).

    abstract：:Recombinant murine MRP14 (mMRP14) was produced in Escherichia coli using the pGEX expression system. The mass of fusion protein, by electrospray ionization-mass spectrometry (ESI/MS), was 39,213 Da which compares well with the theoretical mass (39,210.4 Da). Thrombin digestion of fusion protein was expected at a clone...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.1998.1015

    authors： Raftery MJ,Collinson L,Geczy CL

    更新日期：1999-03-01 00:00:00

  • Affinity purification of ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit epsilon N-methyltransferase.

    abstract：:Ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit epsilon N-methyltransferase (Protein methylase III, Rubisco LSMT, EC 2.1.1.43) catalyzes methylation of the epsilon-amino group of Lys-14 in the large subunit of Rubisco. In this paper, an affinity purification procedure for pea (Pisum sativum L. cv Laxton'...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.1995.1070

    authors： Wang P,Royer M,Houtz RL

    更新日期：1995-08-01 00:00:00

  • Arabidopsis thaliana plants expressing Rift Valley fever virus antigens: Mice exhibit systemic immune responses as the result of oral administration of the transgenic plants.

    abstract：:The zoonotic Rift Valley fever virus affects livestock and humans in Africa and on the Arabian Peninsula. The economic impact of this pathogen due to livestock losses, as well as its relevance to public health, underscores the importance of developing effective and easily distributed vaccines. Vaccines that can be del...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2016.07.003

    authors： Kalbina I,Lagerqvist N,Moiane B,Ahlm C,Andersson S,Strid Å,Falk KI

    更新日期：2016-11-01 00:00:00

  • Expression and purification of the alpha-subunit of eukaryotic initiation factor eIF2: use as a kinase substrate.

    abstract：:The alpha-subunit of eukaryotic initiation factor eIF2 (eIF2alpha) plays an important role in the regulation of mRNA translation through modulation of the interaction of eIF2 and a second initiation factor, eIF2B. The interaction of the two proteins is regulated in vivo by phosphorylation of eIF2alpha at Ser51. In the...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.1998.0863

    authors： Kimball SR,Horetsky RL,Jagus R,Jefferson LS

    更新日期：1998-04-01 00:00:00

  • Heterologous expression and purification of a marine alginate lyase in Escherichia coli.

    abstract：:Alginate lyase digestion is an efficient way to degrade alginate into oligosaccharides, which are useful in various areas including chemistry, pharmacy and food industry. Here we determined the sequence of Vibrio sp. QY102 sourced alginate lyase, and set up its heterologous expression in E. coli. We improved its secre...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2018.09.002

    authors： Sun X,Shen W,Gao Y,Cai M,Zhou M,Zhang Y

    更新日期：2019-01-01 00:00:00

  • Effect of K225 residue to the catalytic efficiency of Kex2 protease.

    abstract：:The gene encoding S. cerevisiae Kex2 protease derivative Kex2-667 (encoding the N-terminal 20th to 667th amino acid residues of Kex2 protease, containing the propeptide, catalytic domain, P domain and Ser/Thr enrichment region) and its 225th amino acid residue mutant K225L were overexpressed in Pichia pastoris. Protea...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2020.105725

    authors： Yang F,Liu L,Liu Y,Li S

    更新日期：2020-12-01 00:00:00

  • Recombinant expression of biologically active rat leptin in Escherichia coli.

    abstract：:Leptin is a 16-kDa nonglycosylated hormone that is produced in mature adipocytes and which acts primarily in the hypothalamus to reduce food intake and body weight. While the rat is a representative laboratory animal model in obesity research, so far recombinant rat leptin was not available. In the present study, rat ...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.2001.1412

    authors： Park JH,Lee HH,Na SY,Ju SK,Lee YJ,Lee MK,Kim KL

    更新日期：2001-06-01 00:00:00

  • Heterologous expression and in vitro assembly of the transmembrane cytochrome b6.

    abstract：:Folding and assembly studies with alpha-helical membrane proteins are often hampered by the absence of high-level expression systems as well as by missing suitable in vitro refolding procedures. Experimental constraints and requirements for heterologous expression and in vitro assembly of cytochrome b6 have been exami...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2007.08.007

    authors： Prodöhl A,Dreher C,Hielscher R,Hellwig P,Schneider D

    更新日期：2007-12-01 00:00:00

  • Molecular cloning, expression, and functional analysis of the copper amine oxidase gene in the endophytic fungus Shiraia sp. Slf14 from Huperzia serrata.

    abstract：:Huperzine A (HupA) is a drug used for the treatment of Alzheimer's disease. However, the biosynthesis of this medicinally important compound is not well understood. The HupA biosynthetic pathway is thought to be initiated by the decarboxylation of lysine to form cadaverine, which is then converted to 5-aminopentanal b...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2016.07.013

    authors： Yang H,Peng S,Zhang Z,Yan R,Wang Y,Zhan J,Zhu D

    更新日期：2016-12-01 00:00:00

  • PDZ affinity chromatography: a general method for affinity purification of proteins based on PDZ domains and their ligands.

    abstract：:PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ∼90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2014.02.015

    authors： Walkup WG 4th,Kennedy MB

    更新日期：2014-06-01 00:00:00

  • First characterization of extremely halophilic 2-deoxy-D-ribose-5-phosphate aldolase.

    abstract：:2-Deoxy-d-ribose-5-phosphate aldolase (DERA) catalyzes the aldol reaction between two aldehydes and is thought to be a potential biocatalyst for the production of a variety of stereo-specific materials. A gene encoding DERA from the extreme halophilic archaeon, Haloarcula japonica, was overexpressed in Escherichia col...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2016.05.009

    authors： Ohshida T,Hayashi J,Satomura T,Kawakami R,Ohshima T,Sakuraba H

    更新日期：2016-10-01 00:00:00

  • Human müllerian inhibiting substance: enhanced purification imparts biochemical stability and restores antiproliferative effects.

    abstract：:Separation of copurifying protease activity from recombinant human Müllerian inhibiting substance (rhMIS) bound to a monoclonal antibody immunoaffinity column by a high-salt wash results in cleaner preparations of rhMIS resistant to cleavage upon storage. In addition, an inhibitor of rhMIS antiproliferative activity i...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/1046-5928(92)90020-w

    authors： Ragin RC,Donahoe PK,Kenneally MK,Ahmad MF,MacLaughlin DT

    更新日期：1992-06-01 00:00:00

  • A four-step, inexpensive protocol for large-scale purification of goat uterine estrogen receptor.

    abstract：:A relatively inexpensive yet highly efficient and extremely rapid procedure has been developed for the isolation and purification of estrogen receptor from the goat uterine cytosol. Greater than 1 mg of purified receptor protein could be obtained from 75 g of uterine tissue within a period of < 24 h, following this pr...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1006/prep.1993.1070

    authors： Zafar A,Thampan RV

    更新日期：1993-12-01 00:00:00

  • Cloning, expression, purification and functional characterization of the oligomerization domain of Bcr-Abl oncoprotein fused to the cytoplasmic transduction peptide.

    abstract：:Protein-based cellular therapeutics have been limited by getting molecules into cells and the fact that many proteins require accurate cellular localization for function. Cytoplasmic transduction peptide (CTP) is a newly designed transduction peptide that carries molecules across the cell membrane with a preference to...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2008.10.023

    authors： Huang SF,Liu DB,Zeng JM,Xiao Q,Luo M,Zhang WP,Tao K,Wen JP,Huang ZG,Feng WL

    更新日期：2009-04-01 00:00:00

  • Purification, characterization, and crystallization of membrane bound Escherichia coli tyrosine kinase.

    abstract：:Escherichia coli tyrosine kinase (Etk) is a membrane bound kinase in gram-negative bacteria that regulates the export of capsular polysaccharides (CPS). The molecular mechanism behind CPS regulation remains unclear, despite access to a crystal structure of the cytoplasmic kinase domain of Etk. In this study, an effici...

    journal_title：Protein expression and purification

    pub_type： 杂志文章

    doi：10.1016/j.pep.2015.08.029

    authors： Chesterman C,Jia Z

    更新日期：2016-09-01 00:00:00