SALT-DEPENDENT AND PROTEIN-CONCENTRATION-DEPENDENT CHANGES IN THE SOLUTION STRUCTURE OF THE DNA-BINDING HISTONE-LIKE PROTEIN, HBSU, FROM BACILLUS-SUBTILIS

The solution structure of the histone-like DNA-binding protein, HBsu, from Bacillus subtilis in 2 mM sodium cacodylate, pH 7.5, is sensitive to the ionic strength of the buffer. This was shown by circular dichroism measurements at different concentrations of sodium chloride and potassium fluoride. The stability of HBsu is also influenced; at HBsu concentrations of about 0.1 mg . ml-1, melting temperatures of 32-degrees-C and 55-degrees-C were found in the absence of potassium fluoride and in the presence of 0.5 M potassium fluoride, respectively, exhibiting drastic ionic-strength-dependent differences in the temperature-induced unfolding of HBsu. Furthermore, at low ionic strength, circular dichroism spectra vary markedly depending on the HBsu concentration in the approximate range 0.2-3 mg . ml-1. Such protein-concentration-dependent differences in the spectra were not observed in the presence of 0.5 M potassium fluoride. Very similar circular dichroism spectra of HBsu and the histone-like DNA-binding protein of Bacillus stearothermophilus (HBst) at high ionic strength, indicate comparable structures of both proteins under these conditions. Estimation of the secondary structure content from the circular dichroism spectra yields data which are in satisfactory agreement with the values obtained from the crystal structure of HBst. Transition temperatures of 45-degrees-C and 61-degrees-C were found in differential scanning calorimetric measurements performed with HBsu in potassium-fluoride-free buffer and in the presence of 0.5 M potassium fluoride, respectively. The thermodynamic data point to the melting of native HBsu dimers into two denatured monomers.