Purification of biotin-binding protein from chicken egg yolk and comparison with avidin.

A simple alternative procedure for the purification in higher yields of the biotin-binding protein from the chicken egg yolk in a ligand-free form is described. The isolated protein was homogeneous by the criteria of polyacrylamide gel electrophoresis, gel filtration chromatography, immuno-double diffusion and immuno-electrophoresis. The protein had an Mr of 72 000 +/- 2000 and was a homotetramer of subunit Mr of 18 000 +/- 1000. It bound [14C]biotin in the molar ratio of 1:4 with an association constant (Ka) of 0.58 X 10(12) M-1. The yolk biotin-binding protein and avidin exhibited qualitatively similar spectral changes on interaction with biotin and p- hydroxyazobenzoic acid, but quantitatively these changes were more pronounced with avidin. Despite the lack of gross immunological cross-reactivity between the two biotin-binders, evidence based on immunological techniques for some degree of common conformational characteristics restricted to or around the ligand-binding sites of the two proteins was adduced. The mixed subunits of the two proteins failed to form hetero-oligomers on reconstitution.