Abstract：
:Post-translational modification of proteins is a dynamic way of generating new protein-protein interaction interfaces that are critical for signaling networks in diverse cellular functions. Purified recombinant proteins frequently lack these signature modifications. Using the tumor suppressor p53 as the model protein, we present here a tethered catalysis approach for the production of acetylated p53 in vivo. P53 is a major tumor suppressor protein that protects the cell from various oncogenic stresses. Upon DNA damage, p53 is stabilized and activated by a plethora of post-translational modifications, including acetylation. Here, we show that constitutively acetylated p53 can be expressed and purified from both yeast and Escherichia coli. This method is highly suitable for studying protein-protein interactions in the conventional yeast two-hybrid screen that requires a constitutively acetylated state of p53. Furthermore, effective production and purification of acetylated p53 from E. coli supports future biochemical and structural characterization. The method described in this work can be applied to other proteins and modifications, and thus has widespread use in the fields of signal transduction and proteomic research.
journal_name
Protein Expr Purifjournal_title
Protein expression and purificationauthors
Acharya A,Xu XJ,Husain-Ponnampalam RD,Hoffmann-Benning S,Kuo MHdoi
10.1016/j.pep.2005.01.015keywords:
subject
Has Abstractpub_date
2005-06-01 00:00:00pages
417-25issue
2eissn
1046-5928issn
1096-0279pii
S1046-5928(05)00025-2journal_volume
41pub_type
杂志文章abstract：:A recombinant Hc fragment of botulinum neurotoxin, serotype B (rBoNTB(Hc)), has been successfully expressed in a Mut+ strain of the methylotrophic yeast Pichia pastoris for use as an antigen in a proposed human vaccine. The fermentation process consisted of batch phase on glycerol, followed by glycerol and methanol fe...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1998.0910
更新日期：1998-08-01 00:00:00
abstract：:Lactobacillus β-galactosidases are mostly heterodimeric proteins, which are encoded by the two overlapping genes, lacL and lacM, and produced in recombinant prokaryotic systems for higher yield. This is the first report on the expression of a heterodimeric β-galactosidase from Lactobacillus crispatus B470 in Pichia pa...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2013.08.019
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abstract：:Human lysosomal beta-hexosaminidase exists in two major forms: the A isoform is composed of both alpha and beta chains, while the B form is a homopolymer of beta chains. Deficiency of beta-hexosaminidase underlies the GM2 gangliosidoses. We have produced active beta-hexosaminidase B in cultured insect (Sf9) cells by i...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/1046-5928(90)90003-h
更新日期：1990-11-01 00:00:00
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
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abstract：:Plasmids pQE-60 and pQE-30 containing 6 x His-tag sequence were used for expression of fragments of HCV structural and non-structural proteins in Escherichia coli (E. coli). The following fragments were used: core (1-98 aa), NS3 (202-482 aa), and tetramer of hypervariable region 1 (HVR1) of E2 protein. The constructed...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.06.011
更新日期：2006-11-01 00:00:00
abstract：:The major active form of human thrombin, alpha-thrombin, was analyzed by hydrophobic interaction high-performance liquid chromatography (HIC-HPLC). The chromatographic system included a polymeric phenyl column and elution was performed by a gradient, 2-0M sodium chloride (5-20 min). Total analysis time was 30 min per ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(02)00596-x
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journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1995.1044
更新日期：1995-06-01 00:00:00
abstract：:Peptide-based affinity tags are commonly used in recombinant production/purification of proteins, and are often preceded or followed by a protease recognition sequence to allow tag removal. We describe a rat monoclonal antibody 2H5 recognizing an undecapeptide tag called "eTev", which contains a recognition sequence f...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2018.03.004
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journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.05.002
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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. The DNA sequence encoding the light chain (catalytic subunit) of human ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(03)00159-1
更新日期：2003-09-01 00:00:00
abstract：:Cyclohexanone monooxygenase (CMO) is a soluble flavoenzyme originally isolated from Acinetobacter spp. which carries out Baeyer-Villiger reactions with cyclic ketone substrates. In the present study we cloned the Acinetobacter CMO gene and modified it for facile purification from heterologous expression systems by inc...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1340
更新日期：2001-02-01 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：:The Human Secretome Project aims to produce and purify all human secreted proteins as full-length. In order to enable this, a robust, gentle and effective purification process is needed, where multiple proteins can be purified in parallel. For this reason, a purification system based on a Protein C-tag and the HPC4 an...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2020.105698
更新日期：2020-11-01 00:00:00
abstract：:Darbepoetin alfa is an engineered and hyperglycosylated analog of recombinant human erythropoietin (EPO) which is used as a drug in treating anemia in patients with chronic kidney failure and cancer. This study desribes the secretory expression of a codon-optimized recombinant form of darbepoetin alfa in Leishmania ta...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.10.013
更新日期：2016-02-01 00:00:00
abstract：:ABCA4 is a member of the A subfamily of ATP-binding cassette transporters that consists of large integral membrane proteins implicated in inherited human diseases. ABCA4 assists in the clearance of N-retinylidene-phosphatidylethanolamine, a potentially toxic by-product of the visual cycle formed in photoreceptor cells...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.02.010
更新日期：2014-05-01 00:00:00
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journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2013.07.014
更新日期：2013-10-01 00:00:00
abstract：:To facilitate studies of multicomponent protein complexes, I have developed an Escherichia coli expression system which coexpresses up to four polypeptides from a single plasmid. The modular nature of the system enables efficient subcloning of a gene into each of the 4 cassettes in the polycistronic expression vector....
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1363
更新日期：2001-02-01 00:00:00
abstract：:Pseudolysin is the extracellular elastase of Pseudomonas aeruginosa and belongs to the thermolysin-like family of metallopeptidases. Pseudolysin has been identified as a robust drug target and a biotechnologically important enzyme in the tanning industry. Previous attempts to purify active pseudolysin from P. aerugino...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.05.005
更新日期：2015-09-01 00:00:00
abstract：:Recombinant human interleukin-5 (rhIL-5) was expressed in baculovirus-infected insect cells and purified to homogeneity from the culture medium in a single chromatographic step. Beginning with a cDNA encoding the full-length precursor form of human IL-5, including the authentic secretory leader sequence, recombinant b...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1995.1009
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abstract：:A new procedure for the large-scale purification of the recombinant thermostable chitinase (Chi40) cloned from Streptomyces thermoviolaceus in various expression vectors in Escherichia coli is described. Chi40 was overproduced in the cytosolic and secreted forms. The cytosolic form (Chi40c) was highly overproduced and...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2001.1490
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abstract：:The gene from Aeromonas veronii bv. sobria encoding the metallo-beta-lactamase ImiS was subcloned into pET-26b, and ImiS was over-expressed in BL21(DE3) Escherichia coli and purified using SP-Sepharose chromatography. This protocol yielded over 5 mg of ImiS per liter of growth culture under optimum conditions. The bio...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.04.017
更新日期：2004-08-01 00:00:00
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
更新日期：2016-10-01 00:00:00
abstract：:While well established in bacterial hosts, the effect of coding sequence variation on protein expression in mammalian systems is poorly characterized outside of viral proteins or proteins from distant phylogenetic families. The potential impact is substantial given the extensive use of mammalian expression systems in ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2007.05.017
更新日期：2007-10-01 00:00:00
abstract：:Fusion proteins expressed in bacteria are often insoluble or inefficiently purified by standard procedures previously reported to work well for the non-fused proteins. We report here a simple but general procedure that can be used to quickly customize and optimize the purification of milligram quantities of most GST f...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(02)00524-7
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abstract：:A mammalian expression vector with features optimized for simple expression and purification of secreted proteins has been developed. This vector was constructed to facilitate X-ray crystallographic studies of cysteine-rich glycoproteins that are difficult to express by other means. Proteins expressed with this vector...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1331
更新日期：2000-12-01 00:00:00
abstract：:A strategy for the purification and cleavage of chimeric recombinant proteins based on a genetically engineered metal-binding peptide and a human renin cleavage site is described. Vectors were constructed to direct the synthesis of chimeric human immunodeficiency virus (HIV) reverse transcriptase (RT) or beta-galactos...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/1046-5928(91)90073-r
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abstract：:Endo-1,4-β-mannanase is an enzyme that can catalyze the random hydrolysis of β-1,4-mannosidic linkages in the main chain of mannans, glucomannans and galactomannans and offers many applications in different biotechnology industries. Purification and kinetic properties of the endo-1,4-β-mannanase from recombinant Esche...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2018.01.009
更新日期：2018-06-01 00:00:00
abstract：:The AP-1 transcription factor is a dimeric protein complex formed primarily between Jun (c-Jun, JunB, JunD) and Fos (c-Fos, FosB, Fra-1, Fra-2) family members. These distinct AP-1 complexes are expressed in many cell types and modulate target gene expression implicated in cell proliferation, differentiation, and stres...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.01.016
更新日期：2008-05-01 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：:Deformylation of the initiator N-formylmethionine does not always proceed to completion for proteins overexpressed in Escherichia coli. To overcome this limitation, the def gene encoding the Escherichia coli peptide deformylase was cloned into the plysS plasmid under the tetracycline (Tc) promoter control. The efficie...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.03.007
更新日期：2004-07-01 00:00:00
