Characterization of the fluorescence of the protamine thynnine and studies of binding to double‐stranded DNA

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Abstract
The protamine thynnine is an arginine‐rich protein approximately 30 amino acids long with a tyrosine in the middle of its sequence. Its fluorescence decay kinetics can be described by a biexponential function with lifetimes of 0.52 and 2.1 ns, with almost equal preexponential factors. The fluorescence quencher CsCl does not affect the short lifetime but shifts the equilibrium between the long and short lifetime toward the short one and reduces the long lifetime. In nature, thynnine is found complexed with chromosomal DNA. In vitro complexes of thynnine with double‐stranded (ds) DNA are stable at physiologic ionic strength but dissociate at high NaCl concentration. This dissociation can be monitored by steady‐state fluorescence. From the salt concentration dependence of the dissociation of the complex of thynnine with ds‐DNA 145 bp long, it can be concluded that only 4 of 21 possible full electrostatic bonds are involved in thynnine‐DNA binding. In addition, the binding constant at 1M NaCl is of the order of 106, indicating a strong nonelectrostatic component in arginine‐DNA binding.