Fluorescence polarization studies of the interactions between protamine and enzymes in aqueous solutions

Fluorescence polarization has been used to study the interaction of dansylated protamine with the enzymes: pepsin, α-chymotrypsin, alkaline phosphatase and invertase. These interactions have been compared with those between dansylated protamine and polyacrylate, or polyvinylsulphate. Each of the various complexes was found to be dissociated by the addition of sodium nitrate and a critical electrolyte concentration (CEC) was determined for each system, to allow assessment of the relative order of binding to the dansylated protamine. This order was: polyvinylsulphate >pepsin >polyacrylate >alkaline phosphatase >α-chymotrypsin. The strength of binding was also assessed by determination of a binding constant, K_a. The values of K_a showed the same relative order of binding as the CEC values. Invertase behaved similarly to the other enzymes, but it was not possible to obtain an unambiguous assessment of the comparative strength of binding. In each case, the stoichiometry of the complex was also determined.