Binding site of the dye in Bromophenol blue-lysozyme complex. Proton magnetic resonance study in aqueous solutions

The forward rate constant for the binding of Bromophenol blue to lysozyme is estimated to be 10(5)M-1 X S-1 at room temperature and ionic strength 0.01, from the line broadening of the 1H-NMR spectrum of Bromophenol blue at 270 MHz. The broadening of the 1H-NMR line corresponding to histidine-15 of lysozyme, in the presence of Bromophenol blue in solution at pH 4.5, suggests that the binding site of Bromophenol blue is not far from this protein residue. The extent of this broadening is pH-dependent and can be interpreted in terms of the average coulombic interaction between the dye and His-15. Since Bromophenol blue is known to inhibit lytic activity of lysozyme towards cell wall substrates, we can say that the region near His-15 may also be important for the efficient catalytic activity of lysozyme.