Abstract：
:Human granulocyte-macrophage colony-stimulating factor (GM-CSF), a hemopoietic growth factor, was produced and secreted from tobacco cell suspensions. The GM-CSF cDNA was carried by a binary vector under the control of the CaMV 35S promoter and the T7 terminator. In addition, a 5'-nontranslated region from the tobacco etch virus (TEV leader sequence) was fused to the N-terminal end of the GM-CSF transgene. For ease of purification, a 6-His tag was added to the 3' end of the GM-CSF cDNA. Addition of the TEV leader sequence increased protein production more than twofold compared to non-TEV controls. Initial batch cultivation studies indicated a maximum of 250 microg/L extracellular and 150 microg/L intracellular GM-CSF. Western blot analysis detected multiple peptides with masses from 14 to 30 kDa in the extracellular medium. The plant-produced GM-CSF was biologically active and could be bound to a nickel affinity matrix, indicating that both the receptor-binding region and the 6-His tag were functional. The batch production of GM-CSF was compared with the production of other recombinant proteins secreted by transformed tobacco cells. The recovery of secreted GM-CSF was increased by the addition of stabilizing proteins and by increasing salt in the growth medium to physiological levels.
journal_name
Protein Expr Purifjournal_title
Protein expression and purificationauthors
James EA,Wang C,Wang Z,Reeves R,Shin JH,Magnuson NS,Lee JMdoi
10.1006/prep.2000.1232keywords:
subject
Has Abstractpub_date
2000-06-01 00:00:00pages
131-8issue
1eissn
1046-5928issn
1096-0279pii
S1046-5928(00)91232-4journal_volume
19pub_type
杂志文章abstract：:In order to obtain bioactive α-bungarotoxin (αBtx) using recombinant protein technique, a codon-optimized synthetic gene was expressed in fusion with the N-terminal 10-His-tag and C-terminal Strep-tag in Escherichia coli. Further optimization through site-directed mutagenesis enabled moderate expression of the protein...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.12.016
更新日期：2015-06-01 00:00:00
abstract：:An efficient expression system for a low-molecular mass xylanase in Escherichia coli has been developed. A gene encoding the mature Bacillus circulans (Bc) xylanase was designed to imitate the frequency of degenerate codons used in E. coli. Appropriate degenerate codons were used to create multiple unique restriction ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1993.1026
更新日期：1993-06-01 00:00:00
abstract：:Protein convertase 1/3 is a serine endoproteinase present in the regulated secretory pathway of endocrine and neuroendocrine cells. It is responsible for the processing of numerous prohormones and proneuropeptides into their biologically active moieties, often following cleavage at pairs of basic residues. The determi...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.06.014
更新日期：2004-10-01 00:00:00
abstract：:Splicing factor proline- and glutamine-rich (SFPQ) is an RNA-binding protein, playing significant roles in gene regulation and subnuclear body formation. Our recent serendipitous discovery showed that SFPQ binds zinc directly and forms an infinite polymer that is induced by zinc binding to the protein. The zinc-induce...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2020.105626
更新日期：2020-07-01 00:00:00
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
更新日期：1998-04-01 00:00:00
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
更新日期：2007-12-01 00:00:00
abstract：:Pyrroline-5-carboxylate reductase (P5CR) catalyzes the reduction of Delta1-pyrroline-5-carboxylate (P5C) to proline with concomitant oxidation of NAD(P)H to NAD(P)(+). The enzymatic cycle between P5C and proline is very important in many physiological and pathological processes. Human P5CR was over-expressed in Escher...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.02.019
更新日期：2006-09-01 00:00:00
abstract：:Enteroaggregative Escherichia coli (EAEC) heat-stable enterotoxin 1 (EAST1) is a 4.1k Da protein originally discovered in EAEC but known to be scattered in other diarrheagenic E. coli as well, possibly causing diarrhea in humans and animals. We report for the first time a method to express and purify EAST1 using the G...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2003.09.008
更新日期：2004-02-01 00:00:00
abstract：:Streptomyces coelicolor is a soil-dwelling bacterium that undergoes an intricate, saprophytic lifecycle. The bacterium takes up exogenous nucleosides for nucleic acid synthesis or use as carbon and energy sources. However, nucleosides must pass through the membrane with the help of transporters. In the present work, t...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.02.004
更新日期：2015-05-01 00:00:00
abstract：:Der p 2, a major allergen derived from the house dust mite Dermatophagoides pteronyssinus, is one of the most clinically relevant allergens worldwide. Recombinant Der p 2 (rDer p 2) is useful in clinical diagnosis and disease-specific immunotherapy. However, previous studies showed that Der p 2 can only be expressed i...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2016.01.012
更新日期：2016-05-01 00:00:00
abstract：:The human melanocortin 4 receptor (MC4r) was successfully expressed in Sf9 cells using the baculovirus infection system. N- and C-terminally His-tagged receptors generated B(max) values of 14 and 23 pmol receptor/mg membrane protein, respectively. The highest expression level obtained with the C-terminally His-tagged ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.06.018
更新日期：2004-10-01 00:00:00
abstract：:Kynurenine 3-monooxygenase (KMO) is an enzyme central to the kynurenine pathway of tryptophan metabolism. KMO has been implicated as a therapeutic target in several disease states, including Huntington's disease. Recombinant human KMO protein production is challenging due to the presence of transmembrane domains, whic...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2013.11.015
更新日期：2014-03-01 00:00:00
abstract：:Pullulanases are well-known starch-debranching enzymes that are widely used for hydrolysis of a-1,6-glycosidic linkages in starch, pullulan, amylopectin, and other oligosaccharides. Escherichia coli is a popular heterologous expression host for generating target enzymes. However, cells have to be disrupted to obtain t...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2018.09.011
更新日期：2019-03-01 00:00:00
abstract：:Human Relaxin 2 is an insulin-related peptide hormone with a mass of 19,084 Da. The mRNA contains a number of arginine codons that are rarely used by Escherichia coli to produce highly expressed proteins. As a result, expressing this recombinant protein in E. coli is problematic. When human Relaxin 2 was expressed in ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.02.016
更新日期：2008-08-01 00:00:00
abstract：:To study the subunit structure and the active site of human immunodeficiency virus reverse transcriptase (RT), the enzyme was expressed in E. coli and purified to homogeneity in large quantities. The recombinant enzyme consists of two major polypeptides of 66,000 and 53,000 Da in equimolar amounts and a minor species ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/1046-5928(92)90005-h
更新日期：1992-08-01 00:00:00
abstract：:The plant protein production system is a platform that can not only reduce production costs but also produce monoclonal antibodies that do not have the risk of residual proteins from the host. However, due to the difference between post-translational processes in plants and animals, there may be a modification in the ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2019.03.004
更新日期：2019-07-01 00:00:00
abstract：:Arginine has been effectively used in various column chromatographies for improving recovery and resolution, and suppressing aggregation. Here, we have tested the effectiveness of arginine as an eluent in dye-affinity column chromatography using Blue-Sepharose, which binds enzymes requiring adenyl-containing cofactors...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.10.005
更新日期：2007-04-01 00:00:00
abstract：:Hendra virus (family Paramyxoviridae) is a negative sense single-stranded RNA virus (NSRV) which has been found to cause disease in humans, horses, and experimentally in other animals, e.g. pigs and cats. Pteropid bats commonly known as flying foxes have been identified as the natural host reservoir. The Hendra virus ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.07.008
更新日期：2015-12-01 00:00:00
abstract：:Infections caused by Acinetobacter baumannii have emerged as a significant clinical problem due to the increase in infections caused by antibiotic resistant strains. A. baumannii OmpA is a highly conserved membrane protein that has multiple roles in interacting with the host during infection, and thus represents an at...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2010.11.019
更新日期：2011-05-01 00:00:00
abstract：:A process for bacterial expression and purification of the recombinant major wasp allergen Antigen 5 (Ves v 5) was developed to produce protein for diagnostic and therapeutic applications for type 1 allergic diseases. Special attention was focused on medium selection, fermentation conditions, and efficient refolding p...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.01.009
更新日期：2006-06-01 00:00:00
abstract：:NIP1, the product of the avirulence gene AvrRrs1 from Rhynchosporium secalis, a fungal pathogen of barley, is a small secreted cysteine-rich protein. This protein is essential for the specific recognition of the fungus by host plants carrying the complementary resistance gene Rrs1. Different heterologous expression sy...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1999.1098
更新日期：1999-10-01 00:00:00
abstract：:Proteins are essential throughout the biological and biomedical sciences and the purification strategies of proteins of interest have advanced over centuries. Elastin-like polypeptides (ELPs) are compound polymers that have recently been highlighted for their sharp and reversible phase transition property when heated ...
journal_title：Protein expression and purification
pub_type： 杂志文章,评审
doi：10.1016/j.pep.2018.09.006
更新日期：2019-01-01 00:00:00
abstract：:The UDP-N-acetylmuramoyl-L-alanine:D-glutamate ligase from Escherichia coli, an enzyme involved in the biosynthesis of the bacterial peptidoglycan monomer unit, was overproduced and purified to homogeneity on a large scale, yielding 4 mg of protein per liter of bacterial culture. Crystals of the complex with the subst...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1997.0850
更新日期：1998-06-01 00:00:00
abstract：:Enteropeptidase (synonym: enterokinase, EC 3.4.21.9) is a heterodimeric serine protease of the intestinal brush border that activates trypsinogen by highly specific cleavage of the trypsinogen activation peptide following the sequence (Asp)(4)-Lys. It has also great biotechnological interest because of the unique subs...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2011.04.005
更新日期：2011-10-01 00:00:00
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
更新日期：2006-03-01 00:00:00
abstract：:Interferon α-2a (IFNA2) is a member of the Type I interferon cytokine family, known for its antiviral and anti-proliferative functions. The role of this family in the innate immune response makes it an attractive candidate for the treatment of many viral and chronic immune-compromised diseases. Recombinant IFNA2 is cl...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.04.010
更新日期：2014-07-01 00:00:00
abstract：:The second enzyme of the naphthalene degradation pathway in Pseudomonas putida G7 is NahB, a dehydrogenase that converts cis-1,2-dihydroxy-1,2-dihydronaphthalene to 1,2-dihydroxynaphthalene. We report the cloning, optimization of expression, purification, kinetic studies and preliminary structural characterization of ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2017.01.005
更新日期：2017-04-01 00:00:00
abstract：:We have recently identified a new family of multidomain oxidoreductase (redox) enzymes, the MICALs, that directly regulate the actin cytoskeletal elements necessary for the morphology, motility, and trajectory of cells. Our genetic assays reveal that Mical is both necessary and sufficient for actin organization and ce...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2016.05.008
更新日期：2016-11-01 00:00:00
abstract：:Arginase (EC 3.5.3.1; L-arginine amidinohydrolase) is a key enzyme of the urea cycle that catalyses the conversion of arginine to ornithine and urea, which is the final cytosolic reaction of urea formation in the mammalian liver. The recombinant strain of the yeast Saccharomyces cerevisiae that is capable of overprodu...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2011.09.001
更新日期：2012-01-01 00:00:00
abstract：:Plants possess very large numbers of biosynthetic cytochrome P450 enzymes. In spite of the importance of these enzymes for the synthesis of bioactive plant secondary metabolites, only two plant P450 structures has been obtained to date. Isoflavone synthase (IFS) is a membrane-associated cytochrome P450 enzyme catalyzi...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2010.11.012
更新日期：2010-12-04 00:00:00
