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#biotin

3 public questions tagged with this topic.

Nucleic acid probe and labelling

Which protein has high affinity for biotin in labeling systems?

Biotin-labeled nucleic acid probes are visualized through exceptionally strong non-covalent protein interactions. Avidin, a tetrameric glycoprotein from egg white, and streptavidin from Streptomyces avidinii bind biotin with a dissociation constant near 10^-15 M, among the strongest known biological affinities. This stability allows efficient capture of biotinylated probes followed by detection using enzyme-conjugated or fluorophore-conjugated avidins. Amplification steps improve sensitivity in blots and histochemistry. Antibodies, alkaline phosphatase, and T4 ligase do not exhibit specific high-affinity biotin binding, making avidin-streptavidin essential for indirect non-isotopic detection systems.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Hybridization

Biotin-tagged proteins are captured by:

Biotin tagging exploits exceptionally strong non-covalent interaction between biotin, also called vitamin H, and streptavidin tetramer, with dissociation constant around 10^-15 M, one of the strongest known in biology. Biotinylated bait or target protein can be efficiently captured on streptavidin coated beads, plates, or sensor chips with minimal non-specific binding due to high specificity. Elution may require denaturing conditions. IgG, TEV, and His-tag resin recognize Fc regions, protease sites, and polyhistidine sequences, respectively, and do not bind biotin. Streptavidin capture preserves protein activity and enables downstream functional assays.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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