Enzymatic Reaction (image will open in a new window)
Avidin is a glycoprotein consisting of four identical subunits. It is a minor constituent of the egg white of reptiles, amphibians, and birds. In the chicken, it is synthesized in the oviduct in response to progesterone and in several tissues in connection with inflammation and tissue damage (Elo et al. 1979, and Gope et al. 1987). It binds the vitamin biotin with high affinity and is therefore thought to be a defense protein against biotin-requiring microbes (Wallén et al. 1995). Avidin exhibits the highest known affinity in nature between a ligand and a protein (Livnah et al. 1993).
In 1916, Bateman first investigated the toxic effect of uncooked egg whites in animals. In 1927, Boas showed that “egg white injury” was caused by a protein in albumin. It was determined that this protein binds biotin forming a “non-digestible” complex that cannot be absorbed from the intestine of animals or from the surrounding medium by microorganisms (Boas 1924, Pennington et al. 1942, Woolley and Longsworth 1942, and Hertz 1946). Eakin et al. first isolated the protein and named it avidin (Eakin et al. 1940a, b, and 1941).
In 1963, Green determined the dissociation constant of the avidin-biotin complex (10-15 M) indicating extreme stability of the complex in vitro (Green 1963a, b). Through the late 1970s and into the early 1980s, the complex formation of the avidin-biotin complex was a useful tool in biochemistry and immunocytochemistry (Guesdon et al. 1979, Bayer and Wilchek 1980 and Hsu et al. 1981). The sequence of chicken avidin was determined by DeLange and Huang in 1971. The cDNA was cloned and sequenced by Gope et al. in 1987, and the gene (avd) was cloned and sequenced by Wallén et al. in 1995.
Recent work with avidin has found its use in a variety of cancer treatments (Lesch et al. 2010). Both avidin and its derivatives continue to be used in the isolation of biotinylated proteins (Bolivar et al. 2008), and it has been incorporated into assays to detect levels of biotin in urine and serum (Zarogiannis et al. 2007).
Avidin is a basic homotetrameric glycoprotein (Green 1964, and DeLange and Huang 1971). The carbohydrate moiety, a polysaccharide composed of four glucosamines and five mannoses, is attached to Asn-17 of each subunit (DeLange and Huang 1971). Each of the identical subunits binds one biotin molecule, and tryptophan is involved in non-covalent binding. Each subunit is folded into an eight-stranded anti-parallel beta-barrel, displaying up and down topology. The biotin binding site is located in the core of the barrel and is built by residues of the barrel itself and the loop of an adjacent subunit (Nardone et al. 1998). Although avidin shares almost identical secondary, tertiary, and quaternary structure with another biotin-binding protein, streptavidin, streptavidin is not glycosylated and is less susceptible to aggregation. Avidin has a higher affinity for unconjugated biotin than streptavidin, but a lower affinity for conjugated biotin (Laitinen et al. 2006).
The best preparations of avidin bind 15.1 µg of D-biotin per milligram of protein, corresponding to 4 moles of biotin bound per mole of protein. The dissociation constant for biotin is approximately 10-15 M, being about 103-106 times higher than that of a typical antigen-antibody interaction (Green 1963a, and Nardone et al. 1998). The biotin-avidin complex is dependent upon media ionic strength, but relatively stable over a wide pH range and to heat.
It is necessary to effect a reversible or irreversible denaturation to free biotin. Guanidinium chloride (6-8 M) at pH 1.5 and autoclaving at 120°C (15 minutes) are examples of sufficient dissociation conditions (Green 1970a, b).
The protein contains a 24 residue signal peptide. The avidin gene (avd) consists of four exons split by three introns (Wallén et al. 1995). Although avidin shares almost identical secondary, tertiary, and quaternary structure with streptavidin, the two proteins show only 30% sequence identity and are thought to be evolutionarily unrelated (Laitinen et al. 2006).
Protein Accession Number: P02701
CATH Classification (v. 3.2.0):
Molecular Weight: 54.7 kDa
Optimal pH: Relatively stable over a wide pH range
Isoelectric Point: ≈10 (Nardone et al. 1998)
Extinction Coefficient: E 1%, 282 = 15.5 (Melamed and Green 1963)