Abstract:
:The fluorescent reporter enhanced Green Fluorescent Protein (EGFP) has been used for assaying a wide range of biological activities ranging from gene expression, or localization of target proteins through to intermolecular interactions. However, over-production of this protein in Escherichia coli has resulted in the presence of inclusion bodies, which complicates recovery of the protein in significant quantities. In this paper, we describe a single-step method for isolating the protein from a Glutathione-S-Transferase (GST) fusion protein, release of the EGFP protein from the fusion was demonstrated using a biotinylated variant of Human Rhinovirus 14 3C protease that we have also constructed. We also suggest the potential uses of the biotinylated protease for bionanotechnology and synthetic biology.
journal_name
Protein Expr Purifjournal_title
Protein expression and purificationauthors
Youell J,Fordham D,Firman Kdoi
10.1016/j.pep.2011.05.003subject
Has Abstractpub_date
2011-10-01 00:00:00pages
258-62issue
2eissn
1046-5928issn
1096-0279pii
S1046-5928(11)00117-3journal_volume
79pub_type
杂志文章abstract::Previously, we have shown that the entire extracellular domain of the granulocyte-colony stimulating factor receptor (sG-CSFr) produced in Chinese hamster ovary (CHO) cells forms a stable complex with its ligand G-CSF, at a stoichiometry of 2:2. A truncated receptor molecule consisting of the cytokine receptor homolog...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1006/prep.1998.0942
更新日期:1998-10-01 00:00:00
abstract::Expression and purification of human beta-secretase (BACE1) in bacteria have been plagued with issues concerning solubility, inhomogeneous N-terminus, and lack of enzymic activity. Several forms of the mature human BACE1 have been expressed in Escherichia coli with different N-terminal extensions and without the C-ter...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2003.12.014
更新日期:2004-04-01 00:00:00
abstract::Human HEK293 cells that stably express the Epstein Barr nuclear antigen 1 (EBNA1) support the episomal replication of plasmids containing the Epstein Barr virus origin of replication (EBV oriP). A 293EBNA (293E) cell line expressing the human corticotropin-releasing hormone receptor subtype I (CRHR1) from an episomal ...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1006/prep.1996.0701
更新日期:1997-04-01 00:00:00
abstract::Granulins (GRNs) are potent growth factors that are upregulated in many aggressive cancers from a wide range of organs. GRNs form tight, disulphide bonded, beta hairpin stacks, making them difficult to express in recombinant form. We recently described Ov-GRN-1, a GRN family member secreted by the carcinogenic liver f...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2011.06.018
更新日期:2011-10-01 00:00:00
abstract::Acetyl-CoA C-acetyltransferase synthase gene (AACT) cDNA, DNA and promoter were cloned from Sanghuangporus baumii. The gene ORF (1260 bp) encoded 419 amino acids. The AACT DNA includes five exons (1-84 bp, 140-513 bp, 570-1027 bp, 1090-1282 bp, 1344-1494 bp) and four introns (85-139 bp, 514-569 bp, 1028-1089 bp, 1283-...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2020.105592
更新日期:2020-06-01 00:00:00
abstract::The limit dextrinase inhibitor (LDI) from barley seeds acts specifically on limit dextrinase (LD), an endogenous starch debranching enzyme. LDI is a 14 kDa hydrophobic protein containing four disulfide bonds and one unpaired thiol group previously found to be either glutathionylated or cysteinylated. It is a member of...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2011.04.009
更新日期:2011-10-01 00:00:00
abstract::Malassezia globosa is pathogenic fungus that causes skin disorders including dandruff in humans. Many yeast cytochrome CYP enzymes are involved in the biosynthesis of sterols and are considered major targets of azole antifungal agents. Here, we report on the expression and characterization of the MGL_0310 gene product...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2015.07.002
更新日期:2015-10-01 00:00:00
abstract::A cDNA clone, lambda GTHP1del, encoding glutathione transferase (GST) P1-1, was isolated from a human K562 erythroleukemia cell line cDNA library. The coding sequence was lacking the codons for the N-terminal 34 amino acids. A DNA segment was designed in order to obtain the missing portion and a structure representing...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1006/prep.1995.1034
更新日期:1995-06-01 00:00:00
abstract::To produce a monoclonal antibody specific to a mouse major histocompatibility complex (MHC) class II protein, we synthesized the complementary DNAs for the heavy and light chains of a monoclonal antibody by PCR amplification. These cDNAs were then introduced separately into tobacco plant cells. After performing Northe...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2007.02.015
更新日期:2007-07-01 00:00:00
abstract::The transcription factor Sp1 is a regulator of TATA-less genes. It belongs to the Cys₂-His₂ zinc finger domain-containing family. A zebrafish cDNA encoding a peptide homologous to mammalian Sp1 was cloned and inserted into a pET43.1a vector and expressed in Escherichia coli Rosetta (DE3) cells as a Nus-His-tag fusion ...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2010.10.010
更新日期:2011-03-01 00:00:00
abstract::A stable mammalian cell line expressing highly active bovine interferon-gamma (BoIFN-γ) was generated using Flp recombinase-mediated integration. This recombinant 293 cell line (B1) efficiently secreted FLAG-tagged BoIFN-γ protein into the culture supernatant, as determined by ELISA and Western blot. The recombinant B...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2014.04.012
更新日期:2014-07-01 00:00:00
abstract::Hydroxypyruvate reductase (HPR), a plant leaf peroxisomal enzyme involved in the glycolate pathway, has been purified in two steps from a crude extract of parsley leaves during the purification of an unrelated ATP-dependent enzyme. HPR, a homogenous side-fraction arising from this purification procedure, was identifie...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1006/prep.1996.0666
更新日期:1997-02-01 00:00:00
abstract::The presence of antibiotic resistance genes in genetically modified bacteria raises a regulatory concern in the production of therapeutic proteins and additionally reduces the number of plasmids available for propagation in a cell. Cre recombinase from bacteriophage P1, involved in Cre/loxP mechanism is one of the wid...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2019.105546
更新日期:2020-03-01 00:00:00
abstract::Fab (fragment that having the antigen binding site) of a monoclonal antibody (mAb) is widely required in biopharmaceutical research and development. At Centocor, two routes of Fab production and purification were used to enable a variety of research and development efforts, particularly, crystallographic studies of an...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2009.04.012
更新日期:2009-10-01 00:00:00
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
更新日期:2004-09-01 00:00:00
abstract::We have successfully expressed and purified active human glycogen synthase-1 (hGYS1). Successful production of the recombinant hGYS1 protein was achieved by co-expression of hGYS1 and rabbit glycogenin (rGYG1) using the MultiBac baculovirus expression system (BEVS). Functional measurements of activity ratios of hGYS1 ...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2013.05.007
更新日期:2013-08-01 00:00:00
abstract::Mammalian target of rapamycin (TOR) controls cell growth and metabolism in response to the availability of nutrients, growth factors, and the cellular energy status. Misregulation of TOR can result in a pathogenic increase or decrease in organ size and in cancer. TOR can be inhibited by binding of a complex of rapamyc...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2008.01.014
更新日期:2008-05-01 00:00:00
abstract::Human interleukin-7 (IL-7) is a member of the interleukin family. Numerous studies have demonstrated IL-7's effect on B- and T-cell development as well as its potential in various clinical applications. Previously, a study reported that IL-7 could be purified from inclusion bodies using a prokaryotic system, however, ...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2008.08.009
更新日期:2009-01-01 00:00:00
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
更新日期:1992-06-01 00:00:00
abstract::The H-NS protein is one of the major constituents of the nucleoid structure that has been implicated in the DNA packaging and in the global regulation of gene expression. The study of this transcriptional regulator is an effort to fight Xylella fastidiosa, a citrus pathogen responsible for a range of economically impo...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/S1046-5928(03)00193-1
更新日期:2003-11-01 00:00:00
abstract::Post-translational modification plays crucial roles in signal transduction in eukaryotic cells. To elucidate the biological function of a protein with a specific post-translational modification, it is necessary to isolate the modified protein. However, it is difficult to incorporate a modified amino acid into a specif...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2007.10.018
更新日期:2008-02-01 00:00:00
abstract::Thymosin α1-thymopentin (Tα1-TP5) fusion peptide has been proved to be an immune regulator based on its higher immunoregulatory activity than Tα1 and TP5. To obtain Tα1-TP5 more effectively and economically, Tα1-TP5 was genetically fused to a self-cleaving intein-chitin binding domain tag for purification via chitin b...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2012.04.013
更新日期:2012-07-01 00:00:00
abstract::Methionine aminopeptidases (MetAPs), ubiquitous enzymes that play an important role in nascent protein maturation, have been recognized as attractive targets for the development of drugs against pathogenic protozoa including Plasmodium spp. Here, we characterized partial biochemical properties of a type I MetAP of Pla...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2015.01.003
更新日期:2015-04-01 00:00:00
abstract::The Ig-binding properties of protein L from Peptostreptococcus magnus and protein G from Streptococcus have been successfully combined through the construction of a novel hybrid protein, consisting of a single Ig-binding domain from each protein. The biophysical and biochemical properties of this construct have been c...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2007.11.007
更新日期:2008-03-01 00:00:00
abstract::Streptococcal protein G has been used extensively for the purification of antibodies using the interaction of the Fc region with protein G. Many antibodies also interact with protein G through a low-affinity binding site for the Fab region. The exploitation of this low-affinity interaction for the purification of Fab'...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/s1046-5928(05)80037-3
更新日期:1992-10-01 00:00:00
abstract::The Gradiflow, a preparative electrophoresis instrument designed to separate molecules on the basis of their size and charge, was used to purify antibody Fab and F(ab')2 fragments. The method described is charge based, utilizing the difference in the pI between the antibody Fab/F(ab')2 fragments and antibody Fc fragme...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/S1046-5928(03)00219-5
更新日期:2003-11-01 00:00:00
abstract::Ricin A chain (RTA) mutants which had been modified by the addition of three lysine residues, three lysines and an alanine, or six histidine residues at the carboxyl terminus were expressed in Escherichia coli. The recombinant proteins were purified to homogeneity by ion-exchange chromatography on CM-Sepharose CL-6B. ...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1006/prep.1995.1087
更新日期:1995-10-01 00:00:00
abstract::Trichoderma reesei tyrosinase TYR2 has been demonstrated to be able to oxidize various phenolic compounds and also peptide and protein bound tyrosine, and thus is of great interest for different biotechnological applications. In order to understand the reaction mechanism of the enzyme it would be essential to solve it...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2007.04.014
更新日期:2007-09-01 00:00:00
abstract::The expression of human lactoferrin in the mammary gland is an attractive approach to diminish its current production cost. Previous attempts to produce lactorferrin in the milk of transgenic animals resulted in very high cost and uncertain results. In this paper, we have directly infused replication-defective adenovi...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2006.11.019
更新日期:2007-05-01 00:00:00
abstract::Fatty acid desaturases are enzymes that introduce double bonds into fatty acyl chains, among which stearoyl-acyl carrier protein desaturase (S-ACP-DES) was widely distributed in the plant kingdom. We cloned the cDNA coding for fab2/ssi2, an S-ACP-DES from Arabidopsis thaliana, into the vector pET30a and heterologously...
journal_title:Protein expression and purification
pub_type: 杂志文章
doi:10.1016/j.pep.2009.08.011
更新日期:2010-02-01 00:00:00