Maltose Binding Protein Fusion
Maltose Binding Protein Fusion - Related Videos
11:19
Novel RNA-Binding Proteins Isolation by the RaPID Methodology
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2016
RNA-protein interactions lie at the heart of many cellular processes. Here, we describe an in vivo method to isolate specific RNA and identify novel proteins that are associated with it. This could shed new light on how RNAs are regulated in the cell.
07:51
Pull-down of Calmodulin-binding Proteins
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Cited by 19 •
2012
Calmodulin (CaM) pull-down assay is an effective way to investigate the interaction of CaM with various proteins. This method uses CaM-sepharose beads for efficient and specific analysis of CaM-binding proteins. This provides an important tool to explore CaM signaling in cellular function.
09:27
Generation of Fluorescent Protein Fusions in Candida Species
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Cited by 6 •
2017
PCR-mediated gene modification can be used to generate fluorescent protein fusions in Candida species, which facilitates visualization and quantitation of yeast cells and proteins. Herein, we present a strategy for constructing a fluorescent protein fusion (Eno1-FP) in Candida parapsilosis.
10:37
Comparing the Affinity of GTPase-binding Proteins using Competition Assays
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Cited by 2 •
2015
This protocol compares the relative affinities of binding partners for Rho-family GTPases, including Rac1. In vivo, Rac1-binding proteins compete for a single binding interface, the conformation of which is dictated by a bound nucleotide. The nucleotide is both important and difficult to control experimentally, due to the high hydrolysis rate.
04:06
Single-Molecule Imaging to Visualize Fusion Protein Condensates Assembly on DNA Curtains
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2025
This video describes a method to visualize the assembly of fusion protein condensates on DNA molecules at specific loci using single-molecule imaging. These condensates are formed due to LCD-LCD and LCD-DBD interactions, which allow for the recruitment of RNA polymerase II and enhanced gene transcription.
FAQs
MBP contributes a stable, soluble structure to the fusion protein, which can support proper folding of the attached target sequence. This effect may reduce aggregation during recombinant production and increase the amount of target protein that remains soluble. The strategy is therefore especially useful when the unfused protein is unstable or difficult to recover in a soluble form.
The MBP portion retains its ability to bind amylose, allowing the fusion protein to associate with amylose resin during affinity chromatography. Because the target is genetically linked to MBP, this interaction provides a way to isolate the fusion from other components of the expression mixture. Purification therefore depends on the tag's binding property rather than on the target's own characteristics.
A protease-cleavable linker places a removable connection between MBP and the target protein. After the fusion has been purified, protease treatment can separate the tag from the target, allowing downstream experiments to use the target without MBP attached. This option is important when the tag could interfere with structural analysis, biochemical measurements, or functional assays.
The workflow begins by genetically joining the target sequence to the MBP sequence in an engineered expression system. After production, the fusion protein is captured through its interaction with amylose resin and isolated by affinity chromatography. If required, a protease-cleavable linker permits MBP removal after purification, producing a preparation of the target protein for subsequent analysis.
This approach is most useful when a target protein is difficult to express in a soluble form or tends to aggregate during production. Improving soluble recovery can make sufficient material available for biochemical studies, structural analysis, antibody production, or functional assays. Its value lies in addressing an expression and purification problem before the protein is applied to those investigations.
Purified target proteins from an MBP fusion workflow can support several kinds of biological investigation. Researchers may examine biochemical properties, pursue structural analysis, generate antibodies, or perform functional assays. In each case, the fusion strategy helps provide an isolated protein preparation, while optional removal of MBP can make the material more suitable for studies requiring the target in a less modified form.