NONCOOPERATIVITY OF BIOTIN BINDING TO TETRAMERIC STREPTAVIDIN

Streptavidin tetramers have been separated according to their biotin c ontent by anion exchange chromatography. Biotin-free and biotin-satura ted streptavidin were coincubated. Streptavidin at intermediate ate li gation levels, i.e., with one, two, or three molecules of bound biotin , accumulates over time. A steady state distribution of ligation level s is reached after 2 days. When biotin was allowed to redistribute sta rting from homogeneous populations containing two molecules of biotin per tetramer, a similar steady state distribution of ligation levels w as observed, thereby demonstrating an equilibrium distribution. Quanti fication of this equilibrium indicates that biotin binds to streptavid in with no cooperativity.