Photobiotin Labeling

Biotin is a small vitamin found in tissue and blood and is synthesized by intestinal bacteria. Biotin functions as a prosthetic group for several carboxylases and as a CO₂ carrier. The molecule consists of a ring system covalently linked to the enzyme by a valerate side chain acting as a flexible arm. Avidin and streptavidin are two proteins that strongly bind biotin. Avidins are tetrameric proteins; each subunit has a molecular mass of 13,000–16,000 Da. Avidin is found in chicken egg whites, and a lower affinity variant is found in the yolk. Streptavidin is found in the fungus Streptomyces avidinii . Although the dissociation constant of avidin for biotin is higher than that of streptavidin for biotin (10-¹⁵ M and 10-¹⁴ M , respectively), the nonspecific adsorption of streptavidin to nucleic acids and negatively charged cell membranes is preferred over avidin. This is due to the low isoelectric point of streptavidin (pI 5–6), compared to pI 10 for avidin. The binding characteristics and stability of avidin—biotin and streptavidin—biotin complexes have been extensively explored and applied to develop numerous methods in immunology and molecular biology.