The catalytic action of enzymes exposed to charged substrates outperforms the activity exerted on their neutral counterparts.

Enzymes perform their catalytic action according to mechanisms featuring exquisite specificity, up to the selection of substrate conformers. However, regardless of this high specificity enzymes are able to deal with a repertoire of substrates, whose conversion into reaction products can occur with markedly different rates. Among the factors affecting the velocity of enzyme-catalyzed reactions, the presence in the substrate of an electrostatic charge could be of importance. Here we report on the kinetic parameters of four enzymes (bovine carbonic anhydrase and α-chymotrypsin, Escherichia coli β-galactosidase, and sweet almond β-glucosidase) determined using a NO-containing charged substrate or its neutral counterpart. Remarkably, all the considered enzymes were found more effective when exposed to the charged substrates, featuring K and k values respectively lower and higher than those determined using the neutral substrates. Furthermore, by means of ultrafiltration experiments we detected the binding of o-nitrophenyl-β-d-galactopyranoside to a multiplicity of sites in E. coli β-galactosidase. Overall, our observations suggest that the unspecific binding of substrate to enzyme surface aids the cycling of subsequent catalytic turnovers.