Abstract：
:The cDNA for the alpha i1 protein that had undergone site-directed mutagenesis to change glycine-2 to alanine was ligated into a baculovirus transfer vector. A recombinant virus was obtained by transfecting Sf9 cells with both the wild-type baculovirus DNA and the transfer vector and screening for recombinant plaques. Infection with the recombinant virus led to a high level of expression of the mutated alpha i1 protein in the soluble fraction of the cell. The protein was purified by ammonium sulfate precipitation, gel filtration, and immobilized dye chromatography. The typical yield was 7.5 mg from two 800-ml cultures. The protein showed immunoreactivity to three different alpha i-specific antibodies. It bound guanosine 5'-(gamma-thio)triphosphate (GTP gamma S) with a stoichiometry of 0.63 to 0.91 mol/mol and with a rate constant (kGTP gamma S) of binding of 0.126 min-1. When GTP gamma S bound, the protein was protected from complete tryptic cleavage. The recombinant protein was able to undergo pertussis toxin-catalyzed ADP ribosylation and bind beta gamma subunits but with a reduced affinity compared to that of alpha transducin. Thus using a recombinant baculovirus, a nonmyristylated G protein alpha subunit was abundantly expressed in Sf9 cells and milligram quantities of a functional protein were easily purified.
journal_name
Protein Expr Purifjournal_title
Protein expression and purificationauthors
Jones TL,Woodard C,Spiegel AMdoi
10.1006/prep.1993.1010subject
Has Abstractpub_date
1993-02-01 00:00:00pages
64-71issue
1eissn
1046-5928issn
1096-0279pii
S1046-5928(83)71010-7journal_volume
4pub_type
杂志文章abstract：:As an insect-selective neurotoxin, scorpion long-chain BjαIT is a promising prospect for insecticidal application; however, the difficulty of obtaining natural BjαIT represents the major obstacle preventing analysis of its insecticidal activity against agricultural insect pests. Here, we screened recombinant Pichia pa...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2018.07.013
更新日期：2018-12-01 00:00:00
abstract：:Lactase deficiency problem discourages many adults from consuming milk as a major source of micro- and macronutrients. Enzymatic hydrolysis of lactose is an ideal solution for this problem but such processing adds significant costs. In this study, a cold active β-galactosidase from Planococcus sp-L4 (bgal) was optimiz...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.09.008
更新日期：2016-09-01 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：:Hepatocyte growth factor (HGF) is a multifunctional protein expressed in a variety of cell types and tissues. Here we describe a novel one-step method to separate and identify HGF, based on a unique interaction between HGF and Cu(II). Conditioned medium (CM) from mouse 3T3-L1 adipocytes which contains HGF or purified ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1996.0047
更新日期：1996-05-01 00:00:00
abstract：:An alpha-amylase has been purified from the thermophilic fungus Scytalidium thermophilum. A ninefold purification was achieved in a single step using fluidized bed chromatography wherein alginate was used as the affinity matrix. There are at least two isoenzymes as shown by concanavalin A (Con A)-agarose column chroma...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1308
更新日期：2000-11-01 00:00:00
abstract：:(S)-Hydroxynitrile lyase (Hnl) from the tropical rubber tree Hevea brasiliensis catalyzes the formation of (S)-cyanohydrins from hydrocyanic acid and aldehydes or ketones. This enzyme accepts aliphatic, aromatic, and heterocyclic carbonyl compounds as substrates and is therefore considered a potent biocatalyst for the...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1997.0765
更新日期：1997-10-01 00:00:00
abstract：:Developmentally regulated G-proteins (DRGs) are a highly conserved family of GTP-binding proteins found in archaea, plants, fungi and animals, indicating important roles in fundamental pathways. Their function is poorly understood, but they have been implicated in cell division, proliferation, and growth, as well as s...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2009.05.009
更新日期：2009-10-01 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：:We examined the feasibility of high-level production of recombinant human prolactin, a multifunctional protein hormone, in insect cells using a baculovirus expression system. The human prolactin cDNA with and without the secretory signal sequence was cloned into pFastBac1 baculovirus vector under the control of polyhe...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1290
更新日期：2000-11-01 00:00:00
abstract：:A method for the simplified, reproducible production of both normal and altered versions of human proinsulin has been developed. A polyhistidine/proinsulin fusion protein was expressed using a prokaryotic expression system and partially purified by affinity chromatography. Disulfide bonds within the polypeptide were f...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1998.1024
更新日期：1999-04-01 00:00:00
abstract：:An improved, simplified method for the purification of recombinant, tagged DnaA proteins is described. The presented protocol allowed us to purify soluble DnaA proteins from two different bacterial species: Helicobacter pylori and Streptomyces coelicolor, but it can most likely also be used for the isolation of DnaA p...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.01.010
更新日期：2006-07-01 00:00:00
abstract：:The HIV-1 Gag precursor protein, Pr55(Gag), is a multi-domain polyprotein that drives HIV-1 assembly. The morphological features of HIV-1 suggested Pr55(Gag) assumes a variety of different conformations during virion assembly and maturation, yet structural determination of HIV-1 Pr55(Gag) has not been possible due to ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.04.013
更新日期：2014-08-01 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：:Dengue virus causes serious diseases affecting people in tropical and sub-tropical regions. The nonstructural (NS) protein 2B is an integral membrane protein and important for the regulation of viral protease NS3, which is significant for virus replication. The NS2B-NS3 complex is an important drug target for treating...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2011.08.008
更新日期：2011-12-01 00:00:00
abstract：:The aim of this study was to establish a method of purifying intact complement factor H (CFH) from human plasma. CFH was isolated from human plasma by polyethylene glycol (PEG) precipitation, following three sequential chromatographic columns, which consisted of l-lysine Sepharose column, Resource Q column and Sephacr...
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：:Androgen receptor (AR)-associated coregulator 70 (ARA70) is a cytoplasmic protein that has been characterized to have the ability to induce AR transcriptional activity in response to androgens and anti-androgens in prostate cancer cells. AR has been shown to have an important role in the progression of prostate cancer...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.11.002
更新日期：2005-02-01 00:00:00
abstract：:Phthalate dioxygenase (PDO), a hexamer with one Rieske-type [2Fe-2S] and one Fe (II)-mononuclear center per monomer, and its reductase (PDR), which contains flavin mononucleotide and a plant-type ferredoxin [2Fe-2S] center, are expressed by Burkholderia cepacia at approximately 30mg of crude PDO and approximately 1mg ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.09.004
更新日期：2007-04-01 00:00:00
abstract：:Structural studies of human peptide hormone somatostatin 14 (SS14) require high amounts of isotopically labelled SS14 to be produced. Here we report a method for effective production of isotopically labelled SS14. SS14 was expressed as a fusion protein with thioredoxin in Escherichia coli. Co-expression of a longer po...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.03.011
更新日期：2014-07-01 00:00:00
abstract：:Dehydrodolichyl diphosphate synthase (DDPPs) catalyzes the sequential condensation of isopentenyl diphosphate with farnesyl diphosphate to synthesize long-chain dehydrodolichyl diphosphate, which serves as a precursor of glycosyl carrier in glycoprotein biosynthesis in eukaryotes. To perform kinetic and structural stu...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2001.1511
更新日期：2001-12-01 00:00:00
abstract：:A hexahistidine tag was fused to the N-terminus of apoaequorin. A suitable vector encoding the fusion protein was constructed and used for transformation of Escherichia coli JM109 cells. Apoaequorin was overexpressed under the control of tac promoter. It was found, however, that most of the protein existed in the form...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(02)00614-9
更新日期：2003-02-01 00:00:00
abstract：:A method was developed for the affinity purification of human complement properdin. The preparation is part of an integrated scheme in which over 20 human plasma proteins can be recovered in a highly purified form. The yield of properdin was 5.9 mg from 3 liters of plasma, amounting to a 28% recovery. The crucial step...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1994.1026
更新日期：1994-04-01 00:00:00
abstract：:l-glutamate oxidase (GLOD), encoded by the gox gene, catalyses the transformation of l-glutamic acid into α-ketoglutaric acid (α-KG). In the present study, Pichia pastoris was used for heterologous production of GLOD following optimization of the gox coding sequence for expression in the yeast host. A series of constr...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2016.09.014
更新日期：2017-01-01 00:00:00
abstract：:A long-lasting recombinant human albumin-linker-erythropoietin (EPO) is a human albumin gene fused to the N-terminal of EPO with a (GGSGG)(n)-repeated linker inserted between albumin and EPO. Albumin-EPO fusion genes were co-transfected with the dhfr gene. Albumin-EPO fusion protein has three kinds of sub-types (IALE,...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2009.07.003
更新日期：2009-12-01 00:00:00
abstract：:Molecular chaperones are integral components of the cellular machinery involved in ensuring correct protein folding and the continued maintenance of protein structure. An understanding of these ubiquitous molecules is key to finding cures to protein misfolding diseases such as Alzheimer's and Creutzfeldt-Jacob disease...
journal_title：Protein expression and purification
pub_type： 杂志文章,评审
doi：10.1016/j.pep.2005.08.005
更新日期：2006-03-01 00:00:00
abstract：:Interferons (IFNs) are a family of pleiotropic cytokines used for the treatment of various viral infections and cancers. The low-cost production of IFNs with high biological value and the discovery of IFNs with improved properties are important for the treatment of these diseases as well as for understanding the physi...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(03)00187-6
更新日期：2003-10-01 00:00:00
abstract：:The 42 kDa cleavage product from the carboxyl end of the Plasmodium falciparum merozoite surface protein 1 (MSP1(42)) is an important blood-stage malaria vaccine target. Several recombinant protein expression systems have been used for production of MSP1(42) including yeast (Saccharomyces cerevisiae and Pichia pastori...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2006.06.018
更新日期：2006-11-01 00:00:00
abstract：:This article reviews the development of immobilized-metal affinity chromatography (IMAC) and describes its most important applications. We provide an overview on the use of IMAC in protein fractionation and proteomics, in protein immobilization and detection, and on some special applications such as purification of im...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2011.08.021
更新日期：2011-09-03 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：:HSPC144 is a newly identified gene in human CD34(+) hematopoietic stem/progenitor cells. In this work, we have expressed and purified the 225-residue protein from Escherichia coli BL21 (DE3) and identified a stable fragment HSPC144-P (residues 44-225) by limited proteolysis method. The HSPC144-P fragment exhibits high...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2005.03.008
更新日期：2005-07-01 00:00:00
abstract：:Stem cell factor (SCF) known as the c-kit ligand is a two disulfide bridge-containing cytokine in the regulation of the development and function of hematopoietic cell lineages and other cells such as mast cells, germ cells, and melanocytes. The secreted soluble form of SCF exists as noncovalently associated homodimer ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.09.015
更新日期：2015-01-01 00:00:00
