Assessing the histidine tautomer fractions in proteins. Test on diisopropylfluorophosphatase, a large all-β protein, from Loligo vulgaris

The importance of histidine tautomerism for an accurate drug/protein structure determination and their relevance for the interaction between biological systems is recognized. Consequently, here we used a recently introduced method to determine the pKa values of ionizable residues and fractions of ionized and tautomeric forms of histidine and acid residues in protein, as a function of pH, to analyze a histidine-rich protein, specifically, to study the accuracy of the prediction of the tautomeric fractions of the imidazole ring of each of the 6 histidine residues of diisopropylfluorophosphatase (pdb id 1E1A), a 314-residue all beta-protein, from Loligo vulgaris. The average tautomeric fractions of the His was compute by using an approach that includes, but is not limited to, molecular dynamics simulations coupled with calculations of the ionization states for all 94 ionizable residues of the protein 1E1A in water at pH 6.5 and 300 K. The results of the analysis on protein 1E1A indicate that: (i) the averaged calculated fractions of histidine tautomers, namely for the N^d1-H and N^e2-H forms, respectively, are in good agreement, within ~11%, with the prediction of an NMR-based methodology; and (ii) the tautomer fractions among the six His residues are very different, mainly because the tautomer preferences are determined by the protein environment. This work provides critical insight about the limitations of continuum dielectric models to reproduce NMR-based tautomeric fractions, and may spur significant progress in our effort to develop fast and accurate methods to determine the forms of the imidazole ring of His in proteins as a function of pH.