CBD Tags for Protein Affinity Purification

Overview

Introduction to CBD tags

The chitin-binding domain (CBD) from Bacillus circulans consists of 51 amino acids, which is commonly used as tags for affinity purification of recombinant proteins in combination with self-splicing inteins in bacterial systems. Following affinity selection of the fusion protein on a chitin matrix, the intein undergoes specific cleavage by a thiol reagent or pH and temperature shift which releases the target protein from the chitin-bound tag.

Fig 1. An illustration of the chemistry involved in purification using the intein/chitin protein tag method. (Methods Enzymol, 2015)

View All Affinity Tags for Protein Purification

CBD tags for protein affinity purification

The intein-chitin binding domain (intein-CBD) tag is a combination of a protein self-splicing element (intein) with a chitin-binding domain, and allows for the purification of a native recombinant protein without need for a protease. Intein-CBD E. coli expression vectors were designed in 1997. Commercially available vectors provide for intein-CBD expression on the N-terminus, C-terminus, or both termini of a heterologous protein of interest.

CBD fusion proteins are purified by affinity chromatography on chitin resin. Nonspecific binding to chitin resin can be reduced by a stringent wash including high salt or detergent (2 M NaCl, 0.5% Triton X). Chitin affinity chromatography is not amenable to denaturing reagents such as urea or guanidine hydrochloride. Low concentrations of reducing agents (<1 mM dithiothreitol or <5 mM 2-mercaptoethanol) may be used during purification, but higher concentrations will prematurely activate the intein self-cleavage reaction. Low concentrations of nonionic detergents (i.e., 0.2% Triton-X or Tween 20) may also be used depending on the nature of the fusion protein.

References

Zhao X, Li G, Liang S. Several affinity tags commonly used in chromatographic purification[J]. J Anal Methods Chem. December, 2013.
Mitchell, S. & Lorsch, J. Protein Affinity Purification using Intein/Chitin Binding Protein Tags. Laboratory Methods in Enzymology: Protein Part D 111-125 (2015).
Michael J. Coolbaugh and David W. Wood. Purification of E. coli Proteins Using a Self-Cleaving Chitin- Binding Affinity Tag.

Reagent Solutions

CBD Antibodies

Target	Cat. No.	Product Name	Host	Isotype	Application
CBD	DPAB16941	Anti-Chitin Binding Domain polyclonal antibody	Rabbit	IgG	WB	Inquiry
CBD	CABT-L472M	Mouse Anti-Chitin-binding domain Monoclonal Antibody, clone 2D2	Mouse	IgG1	WB, ELISA	Inquiry

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