Biomaterial Database

Expression and purification of maltose-binding protein fusions. 2001

P Riggs

New England Biolabs, Beverly, Massachusetts, USA.

This unit describes the procedure for subcloning the sequence encoding the protein of interest into an maltose-binding protein (MBP) vector, and expressing and purifying the fusion protein from the cytoplasm. MBP vectors include a sequence that encodes the four-amino-acid recognition site for the specific protease factor Xa. The site is placed so it can be used to separate the protein of interest from MBP after affinity purification. A support protocol provides a pilot experiment for analyzing the solubility, affinity for the amylose resin, and export of a particular fusion protein. An alternate protocol gives instructions for purifying a fusion protein from the periplasm for fusions that are made in the signal sequence vector and are exported. Additional support protocols detail two different chromatographic methods for separating the protein of interest from MBP after factor Xa cleavage.

UI	MeSH Term	Description	Entries
D011993	Recombinant Fusion Proteins	Recombinant proteins produced by the GENETIC TRANSLATION of fused genes formed by the combination of NUCLEIC ACID REGULATORY SEQUENCES of one or more genes with the protein coding sequences of one or more genes.	Fusion Proteins, Recombinant,Recombinant Chimeric Protein,Recombinant Fusion Protein,Recombinant Hybrid Protein,Chimeric Proteins, Recombinant,Hybrid Proteins, Recombinant,Recombinant Chimeric Proteins,Recombinant Hybrid Proteins,Chimeric Protein, Recombinant,Fusion Protein, Recombinant,Hybrid Protein, Recombinant,Protein, Recombinant Chimeric,Protein, Recombinant Fusion,Protein, Recombinant Hybrid,Proteins, Recombinant Chimeric,Proteins, Recombinant Fusion,Proteins, Recombinant Hybrid
D002352	Carrier Proteins	Proteins that bind or transport specific substances in the blood, within the cell, or across cell membranes.	Binding Proteins,Carrier Protein,Transport Protein,Transport Proteins,Binding Protein,Protein, Carrier,Proteins, Carrier
D002846	Chromatography, Affinity	A chromatographic technique that utilizes the ability of biological molecules, often ANTIBODIES, to bind to certain ligands specifically and reversibly. It is used in protein biochemistry. (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed)	Chromatography, Bioaffinity,Immunochromatography,Affinity Chromatography,Bioaffinity Chromatography
D002852	Chromatography, Ion Exchange	Separation technique in which the stationary phase consists of ion exchange resins. The resins contain loosely held small ions that easily exchange places with other small ions of like charge present in solutions washed over the resins.	Chromatography, Ion-Exchange,Ion-Exchange Chromatography,Chromatographies, Ion Exchange,Chromatographies, Ion-Exchange,Ion Exchange Chromatographies,Ion Exchange Chromatography,Ion-Exchange Chromatographies
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005822	Genetic Vectors	DNA molecules capable of autonomous replication within a host cell and into which other DNA sequences can be inserted and thus amplified. Many are derived from PLASMIDS; BACTERIOPHAGES; or VIRUSES. They are used for transporting foreign genes into recipient cells. Genetic vectors possess a functional replicator site and contain GENETIC MARKERS to facilitate their selective recognition.	Cloning Vectors,Shuttle Vectors,Vectors, Genetic,Cloning Vector,Genetic Vector,Shuttle Vector,Vector, Cloning,Vector, Genetic,Vector, Shuttle,Vectors, Cloning,Vectors, Shuttle
D058619	Maltose-Binding Proteins	Periplasmic proteins that bind MALTOSE and maltodextrin. They take part in the maltose transport system of BACTERIA.	Maltose-Binding Protein,Maltose Binding Protein,Maltose Binding Proteins
D019897	Periplasm	The space between the inner and outer membranes of a cell that is shared with the cell wall.	Periplasmic Space,Periplasmic Spaces,Periplasms,Space, Periplasmic,Spaces, Periplasmic
D029968	Escherichia coli Proteins	Proteins obtained from ESCHERICHIA COLI.	E coli Proteins
D033902	Periplasmic Binding Proteins	Periplasmic proteins that scavenge or sense diverse nutrients. In the bacterial environment they usually couple to transporters or chemotaxis receptors on the inner bacterial membrane.	Periplasmic Binding Protein,Binding Protein, Periplasmic,Binding Proteins, Periplasmic,Protein, Periplasmic Binding