Related ArticlesExpression screening, protein purification and NMR analysis of human protein domains for structural genomics.
J Struct Funct Genomics. 2004;5(1-2):119-31
Authors: Folkers GE, van Buuren BN, Kaptein R
Structural genomics, the determination of protein structures on a genome-wide scale, is still in its infancy for eukaryotes due to the number and size of their genes. Low protein expression and solubility of eukaryotic geneproducts are the major bottlenecks in high-throughput (HTP) recombinant protein production with the E. coli expression systems. To circumvent this problem we decided to focus on separate protein domains. We describe here a fast microtiterplate based, expression and solubility screening procedure, using a combination of in vitro and in vivo expression, and purification with nickel-NTA magnetic beads. All steps are optimized for automatic HTP processing using a liquid handling station. Furthermore, large-scale expression and protein purification conditions are optimized, permitting the purification of 24 protein samples per week. We further show that results obtained from the expression screening can be extrapolated to the production of protein samples for NMR. Starting with 81 cloned human protein domains, in vivo expression was detected in 54 cases, and from 28 of those milligrams of protein were purified. An informative HSQC spectrum was recorded for 18 proteins (22%), half of which were indicative of a folded protein. The success rate and quality of the HSQC spectra suggest that the domain approach holds promise for human proteins.
Expression, purification, and reconstitution of the transmembrane domain of the human amyloid precursor protein for NMR studies.
Expression, purification, and reconstitution of the transmembrane domain of the human amyloid precursor protein for NMR studies.
Expression, purification, and reconstitution of the transmembrane domain of the human amyloid precursor protein for NMR studies.
Protein Expr Purif. 2011 Aug 31;
Authors: Chen W, Gamache E, Richardson D, Du Z, Wang C
Abstract
Alzheimer's disease (AD) is the most common type of dementia in elderly people. Senile plaques, a pathologic hallmark of AD, are composed of amyloid ? peptide (A?). A? aggregation produces...
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Expression and purification of myristoylated matrix protein of Mason-Pfizer monkey virus for NMR and MS measurements.
Expression and purification of myristoylated matrix protein of Mason-Pfizer monkey virus for NMR and MS measurements.
Expression and purification of myristoylated matrix protein of Mason-Pfizer monkey virus for NMR and MS measurements.
Protein Expr Purif. 2011 May 24;
Authors: Prchal J, Junkova P, Strmiskova M, Lipov J, Hynek R, Ruml T, Hrabal R
Matrix proteins play multiple roles both in early and late stages of the viral replication cycle. Their N-terminal myristoylation is important for interaction with the host cell membrane during virus...
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Expression and purification of (15)N- and (13)C-isotope labeled 40-residue human Alzheimer's ?-amyloid peptide for NMR-based structural analysis.
Expression and purification of (15)N- and (13)C-isotope labeled 40-residue human Alzheimer's ?-amyloid peptide for NMR-based structural analysis.
Expression and purification of (15)N- and (13)C-isotope labeled 40-residue human Alzheimer's ?-amyloid peptide for NMR-based structural analysis.
Protein Expr Purif. 2011 May 27;
Authors: Long F, Cho W, Ishii Y
Amyloid fibrils of Alzheimer's ?-amyloid peptide (A?) are a primary component of amyloid plaques, a hallmark of Alzheimer's disease (AD). Enormous attention has been given to the structural...
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Bacterial expression, purification, and model membrane reconstitution of the transmembrane and cytoplasmic domains of the human APP binding protein LR11/SorLA for NMR studies.
Bacterial expression, purification, and model membrane reconstitution of the transmembrane and cytoplasmic domains of the human APP binding protein LR11/SorLA for NMR studies.
Bacterial expression, purification, and model membrane reconstitution of the transmembrane and cytoplasmic domains of the human APP binding protein LR11/SorLA for NMR studies.
Protein Expr Purif. 2011 Feb 11;
Authors: Wang X, Gill Jr RL, Zhu Q, Tian F
LR11 (SorLA) is a recently identified neuronal protein that interacts with amyloid precursor protein (APP), a central player...
Expression, Purification, Detergent Screening and Solution NMR Backbone Assignment of
Expression, Purification, Detergent Screening and Solution NMR Backbone Assignment of the Human Potassium Channel Accessory Subunit MiRP1.
Expression, Purification, Detergent Screening and Solution NMR Backbone Assignment of the Human Potassium Channel Accessory Subunit MiRP1.
Protein Expr Purif. 2010 Nov 15;
Authors: Chen L, Lai C, Lai J, Tian C
MiRP1 (MinK related protein 1) is a membrane protein in the KCNE family. It can associate with and modulate various voltage gated potassium channels. Mutations in human MiRP1 have been found to cause many...
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[NMR paper] Design of an expression system for detecting folded protein domains and mapping macro
Design of an expression system for detecting folded protein domains and mapping macromolecular interactions by NMR.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_FREE_120x27.gif http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www.pubmedcentral.nih.gov-corehtml-pmc-pmcgifs-pubmed-pmc.gif Related Articles Design of an expression system for detecting folded protein domains and mapping macromolecular interactions by NMR.
Protein Sci. 1997 Nov;6(11):2359-64
Authors: ...
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[NMR paper] Human nucleotide excision repair protein XPA: expression and NMR backbone assignments
Human nucleotide excision repair protein XPA: expression and NMR backbone assignments of the 14.7 kDa minimal damaged DNA binding domain (Met98-Phe219).
Related Articles Human nucleotide excision repair protein XPA: expression and NMR backbone assignments of the 14.7 kDa minimal damaged DNA binding domain (Met98-Phe219).
J Biomol NMR. 1997 Oct;10(3):313-4
Authors: Buchko GW, Ni S, Thrall BD, Kennedy MA
Expression screening, protein purification and NMR analysis of human protein domains
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