Thermo Scientific EZ-Link Amine-PEG4-Desthiobiotin is a pegylated amine compound whose biotin-like group is elutable from streptavidin, making it ideal for carbodiimide (EDC) conjugation to carboxylates for labeling and purification.
This desthiobiotin compound is an amine-modified analog of biotin that contains a 4-unit polyethylene glycol (PEG) spacer arm. Desthiobiotinylation is accomplished by conjugating the primary amine to carboxyl groups on surface materials, peptides or proteins using the carbodiimide crosslinker EDC (Part No. 22980). EDC activates carboxyl groups to bind to the –NH2 group of the amino-biotin, forming an amide bond. The desthiobiotin tag binds to streptavidin and other biotin-binding proteins with high specificity yet readily elutes with mild conditions (i.e., by competitive displacement with regular, free biotin). As such, this reagent is a useful alternative to Amine-PEG3-Biotin (Part No. 21347) for avidin-biotin techniques in which nondenaturing elution of the labeled proteins is desired. The No-Weigh Format (5 x 1mg resealable vials) eliminates the difficulties associated with weighing small quantities of reagent and maximizes protection of unused reagent from degradation.
- Desthiobiotin – analog of biotin allows easy elution from streptavidin, an ideal feature for affinity purification applications
- Amine-activated – primary amine can be crosslinked to proteins and material surfaces using EDC and other crosslinkers
- Pegylated – polyethylene glycol (PEG) groups in spacer arm enhances water solubility of biotinylated molecules
|Chemical structure of Amine-PEG4-Desthiobiotin. This reagent is often conjugated to carboxyl groups using the crosslinker EDC. For more information, see our review of Carbodiimide Reaction Chemistry.
Properties of Amine-PEG4-Desthiobiotin. We manufacture our reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.
|Spacer arm length
|Mass added to target
Desthiobiotin vs. Biotin:
Desthiobiotin is a single-ring, sulfur-free analog of biotin that binds to streptavidin with nearly equal specificity but less affinity than biotin (1/Kd = 10^11 vs. 10^15M, respectively)[1-2]. Consequently, desthiobiotinylated bait proteins and their interacting partners can be eluted readily and specifically from streptavidin affinity resin using mild conditions based on competitive displacement with free biotin. For pull-down assay experiments with biological samples, this soft-release characteristic of desthiobiotin also helps to minimize co-purification of endogenous biotinylated molecules, which remain bound to streptavidin upon elution of the target protein complexes with free biotin. The modified avidin-biotin affinity system also eliminates the need to use harsh elution conditions that might disassociate complexes and/or damage the target protein or cell. Desthiobiotin-based techniques are ideal when using native or recombinant proteins that are not expressed with a fusion tag and when isolating captured proteins under native conditions, such as targeting intact cells or cell surface proteins.
Labeling with Amine Compounds:
Amino-compounds can be conjugated to functional groups of proteins and other molecules in a variety of ways. The most common method is to crosslink the terminal primary amine to carboxyl groups using EDC carbodiimide crosslinker chemistry. Carboxyl groups (-COOH) occur in aspartate or glutamate residues and the carboxy-terminus of polypeptides. When activated with EDC (Part No. 22890), carboxylates react with amino (–NH2) groups to form amide bonds. Carboxylate molecules and surface materials can be pre-activated using EDC with Sulfo-NHS (Part No. 24510) for subsequent reaction to primary amines (see NHS-ester Chemistry).
- Hirsch, J., et al. (2002). Easily reversible desthiobiotin binding to streptavidin, avidin, and other biotin-binding proteins: uses for protein labeling, detection, and isolation. Analytical Biochemistry 308: 343-357.
- Hofmann, K., et al. (1982). Avidin binding of carboxyl-substituted biotin and analogues. Biochemistry 21: 978-984.
Review of Biotinylation Methods and Applications
Chemistry of Crosslinking (and Labeling Reagents)
Desthiobiotin Product Selection Guide
All Biotinylation Reagents
Avidin, Streptavidin, NeutrAvidin Affinity Resins