Elucidating the Nature of the Streptomyces plicatus β-Hexosaminidase-Bound Intermediate Using ab initio Molecular Dynamics Simulations

By using all-atom ab initio molecular dynamics simulations, the solution pK(a) of the oxazolinium ion intermediate formed during the Streptomyces plicatus beta-hexosaminidase (SpHex)-catalyzed hydrolysis of beta-D-N-acetylglucosaminides is estimated as pK(a) = 7.7. The structure and protonation state of the enzyme-bound intermediate have also been investigated, using hybrid QM/MM methods. The protonation state and conformational properties of the enzyme bound intermediate are found to be sensitive to the protonation state of a number of ionisable residues (other than the aspartate-glutamate catalytic dyad) suggesting that the microscopic electrostatic environment of SpHex not only perturbs the relative magnitudes of the pK(a) values of the Asp side chain carboxylate and oxazolinium ion but also that SpHex binds its intermediate in a distorted conformation with respect to its ground-state conformation in solution.