A spectroscopic investigation of S-trifluoroethylthiopapain. An investigation of the active site of papain.

The pH dependence of the 19F chemical shift and the fluorescence spectrum of S-2,2,2-trifluoro-1,1-dideuteroethyl-thio-paapain are analysed in terms of dependence on the ionisation of aspartic-acid-158 and histidine-159. The 19F probe causes negative cooperativity between these groups, and does not detected any ionisation at high pH. The intermediate chemical exchange rates for the ionisation of aspartic-acid-158 and histidine-159 allow the approxmate rate constants for proton transfer to be calculated. The rather low rate constants are explained in terms of the hydrophobicity of the active-site region and the net positive charge on the enzyme resulting from its high isoelectric point.