Abstract：
:Soluble oligomers and fibrillar deposits of amyloid beta (Abeta) are key agents of Alzheimer's disease pathogenesis. However, the mechanism of amyloid aggregation and its interaction with live cells still remain unclear requiring the preparation of large amounts of pure and different Abeta peptides. Here we describe an Escherichia coli expression system using a fusion protein to obtain either Abeta(1-40) or Abeta(1-42) by essentially the same procedure. The fusion protein uses a His-tagged intestinal fatty acid binding protein (IFABP) followed by a six-glycine linker and a Factor Xa cleavage site before the Abeta. The advantages of this system are that the fusion protein can be expressed in large amounts, that the fusion partner, IFABP, has been well characterized in terms of folding, that Abeta or mutated Abeta peptides can be obtained without any extra residues attached to the N-terminus and that the system can be used to incorporate fluorine-labeled amino acids. The incorporation of fluorine-labeled amino acids using auxotrophic strains is a useful NMR probe of side chain behavior. We obtain final yields of 4 and 3mg/L of culture for Abeta(1-40) and Abeta(1-42), respectively.
journal_name
Protein Expr Purifjournal_title
Protein expression and purificationauthors
Garai K,Crick SL,Mustafi SM,Frieden Cdoi
10.1016/j.pep.2009.02.009subject
Has Abstractpub_date
2009-07-01 00:00:00pages
107-12issue
1eissn
1046-5928issn
1096-0279pii
S1046-5928(09)00039-4journal_volume
66pub_type
杂志文章abstract：:Quinolinate synthetase catalyzes the second step of the de novo biosynthetic pathway of pyridine nucleotide formation. In particular, quinolinate synthetase is involved in the condensation of dihydroxyacetone phosphate and iminoaspartate to form quinolinic acid. To study the mechanism of action, the specificity of the...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1999.1153
更新日期：2000-02-01 00:00:00
abstract：:A baculovirus vector system that expresses cloned DNA sequences as glutathione S-transferase fusion proteins was developed. This system was used to express and purify the lymphocyte-specific tyrosine kinase p56lck. This recombinant p56lck was purified to homogeneity in a single chromatography step using glutathione re...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1993.1051
更新日期：1993-10-01 00:00:00
abstract：:Human extracellular superoxide dismutase (hEC-SOD) is an enzyme that scavenges reactive oxygen species (ROS). Because of its antioxidant activity, hEC-SOD has been used as a therapeutic protein to treat skin disease and arthritis in mammalian systems. In this study, codon-optimized hEC-SOD was expressed in tobacco (Ni...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2015.11.014
更新日期：2016-03-01 00:00:00
abstract：:The iron-sulfur protein subunit, known as the Rieske protein, is one of the central components of the cytochrome b(6)f complex residing in chloroplast and cyanobacterial thylakoid membranes. We have constructed plasmids for overexpression in Escherichia coli of full-length and truncated Rieske (PetC) proteins from the...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(03)00016-0
更新日期：2003-05-01 00:00:00
abstract：:BTK-2, a 32 residue scorpion toxin initially identified in the venom of red Indian scorpion Mesobuthus tamulus was cloned, overexpressed and purified using Cytochrome b(5) fusion protein system developed in our laboratory. The synthetic gene coding for the peptide was designed taking into account optimal codon usage b...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2009.10.017
更新日期：2010-04-01 00:00:00
abstract：:In order to produce sufficient quantities of fibroblast growth factor-saporin (rFGF-2-SAP) mitotoxin for preclinical evaluation in models of diseases such as cancer and restenosis, we have undertaken the large-scale expression, purification, and characterization of the recombinant molecule. The fusion gene encoding rF...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1996.0079
更新日期：1996-08-01 00:00:00
abstract：:Expression of recombinant proteins in bacterial or eukaryotic systems often results in aggregation rendering them unavailable for biochemical or structural studies. Protein aggregation is a costly problem for biomedical research. It forces research laboratories and the biomedical industry to search for alternative, mo...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2012.10.007
更新日期：2013-02-01 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：:The recombinant human malonyl-CoA decarboxylase (hMCD) was overexpressed in Escherichia coli with and without the first 39 N-terminal amino acids via a cleavable MBP-fusion construct. Proteolytic digestion using genenase I to remove the MBP-fusion tag was optimized for both the full length and truncated hMCD. The apo-...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2003.11.023
更新日期：2004-04-01 00:00:00
abstract：:The coding region for arginyl-tRNA synthetase from jack bean (Canavalia ensiformis) has been sequenced and cloned into the bacterial expression vector pET32a. Transformation of BL21 cells and induction with IPTG results in the high level expression of the protein fused N-terminally with thioredoxin and bearing a His-t...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.05.008
更新日期：2008-10-01 00:00:00
abstract：:A simple and efficient method for hyperexpression in Escherichia coli and purification of capsid protein, p24, of human immunodeficiency virus type 1 (HIV-1) of both B- and C-subtypes is described. DNA-encoding p24 of C-subtype was cloned from C-subtype gag sequence which was obtained by PCR amplification using DNA ex...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2000.1266
更新日期：2000-08-01 00:00:00
abstract：:PLD's (Phospholipases D) are ubiquitously expressed proteins involved in many transphosphatidylation reactions. They have a bi-lobed structure composed by two similar domains which at their interface reconstitute the catalytic site through the association of the two conserved HxKx(4)Dx(6)GSxN motifs. PLD1 interacts wi...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.02.012
更新日期：2008-06-01 00:00:00
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
更新日期：2007-07-01 00:00:00
abstract：:Protein insolubility often poses a significant problem during purification protocols and in enzyme assays, especially for eukaryotic proteins expressed in a recombinant bacterial system. The limited solubility of replication factor C (RFC), the clamp loader complex from Saccharomyces cerevisiae, has been previously do...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2012.03.010
更新日期：2012-06-01 00:00:00
abstract：:An expression library was generated from a partial NcoI and HindIII digest of genomic DNA from the thermophilic bacterium, Bacillus stearothermophilus P1. The DNA fragments were cloned into the expression vector pQE-60 and transformed into Escherichia coli M15[EP4]. Sequence analysis of a lipase gene showed an open re...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2001.1456
更新日期：2001-08-01 00:00:00
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
更新日期：2006-02-01 00:00:00
abstract：:Vibriolysin, an extracellular protease of Vibrio proteolyticus, is synthesized as a preproenzyme. The N-terminal propeptide functions as an intramolecular chaperone and an inhibitor of the mature enzyme. Extracellular production of recombinant vibriolysin has been achieved in Bacillus subtilis, but not in Escherichia ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.08.001
更新日期：2008-12-01 00:00:00
abstract：:High affinity interaction between S-protein and S-peptide fragments of bovine pancreatic RNase A has been recently used for construction of molecular vehicles for targeted drug delivery. The vehicle is assembled as a complex of drug carrier conjugated S-protein with S-peptide-tagged targeting protein. To avoid random ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/s1046-5928(02)00546-6
更新日期：2002-12-01 00:00:00
abstract：:Glycolate oxidase is a flavin-dependent enzyme in the photorespiratory pathway in plants. Here we report the heterologous expression of glycolate oxidase in Escherichia coli and an isolation procedure which results in 4 mg pure protein per gram cell paste in only 1.5 days. This corresponds to a more than 50-fold impro...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.1996.0103
更新日期：1996-11-01 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：: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：: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 prolactin was expressed in insect culture cells by recombinant baculoviruses carrying prolactin gene cDNA placed under the transcriptional control of polyhedrin gene promoter of Autographa californica nuclear polyhedrosis virus. Preliminary results of recombinant human prolactin expression as extracellular as we...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1006/prep.2001.1419
更新日期：2001-07-01 00:00:00
abstract：:The ELR-CXC chemokines are important to neutrophil inflammation in many acute and chronic diseases. Among them, CXCL8 (interleukin-8, IL-8), binds to both the CXCR1 and CXCR2 receptors with high affinity and the expression levels of CXCL8 are elevated in many inflammatory diseases. Recently, an analogue of human CXCL8...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2008.04.007
更新日期：2008-09-01 00:00:00
abstract：:Vascular leak syndrome (VLS) is the major dose-limiting toxicity of immunotoxin therapy. In our previous study, a modified PE38KDEL, denoted PE38KDELKQK, was engineered to eliminate VLS. The PE38KDELKQK-based immunotoxin has been proved to retain potent anti-tumor activity but with a remarkable attenuation in VLS. In ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2007.09.009
更新日期：2008-03-01 00:00:00
abstract：:The major immunodominant integral outer membrane protein C (OmpC) from Salmonella typhi Ty21a was overexpressed, without the signal peptide, in Escherichia coli. The protein aggregates as inclusion bodies (IBs) in the cytoplasm. OmpC from IBs was solubilized with 4 M urea and refolded. This involved rapid dilution of ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2004.12.023
更新日期：2005-03-01 00:00:00
abstract：:Formation of biominerals often involves specific proteins that modulate the process of matrix assembly, nucleation, and crystal growth. AP7 is an aragonite-associated protein of 7 kDa and is intrinsically disordered. The structural disorder of AP7 makes it very difficult to express in Escherchiacoli. In this work, we ...
journal_title：Protein expression and purification
pub_type： 杂志文章
doi：10.1016/j.pep.2014.05.002
更新日期：2014-08-01 00:00:00
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
更新日期：2014-06-01 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：:Over-expression of heterologous proteins in Escherichia coli is commonly hindered by the formation of inclusion bodies. Nevertheless, refolding of proteins in vitro has become an essential requirement in the development of structural genomics (proteomics) and as a means of recovering functional proteins from inclusion...
journal_title：Protein expression and purification
pub_type： 杂志文章,评审
doi：10.1016/j.pep.2007.05.014
更新日期：2007-10-01 00:00:00
