Not only an inhibitor: Trehalose enhances the catalytic action exerted on oxaloacetate by rabbit lactate dehydrogenase.

Trehalose is an osmolyte featuring a prominent competence in stabilizing proteins and enzymes. In particular, in the presence of this peculiar disaccharide, quite a number of enzymes show improved stability against thermal denaturation and an enhanced ability to withstand exposure to freezing-thawing cycles. Moreover, it was recently reported that trehalose counteracts the acid-induced dissociation of oligomeric protein complexes. Concerning the catalytic action of enzymes, the addition of trehalose to assay mixtures was found to decrease the values of both K and k, with the decrease in reaction velocity related to the increase in viscosity induced by the disaccharide. Here, we show that trehalose does not necessarily perform as a negative effector on reaction velocity. Using tetrameric rabbit muscle lactate dehydrogenase (rbLDH) as a model system, we report that trehalose does highly stimulate the catalytic action of this enzyme at the expense of oxaloacetate. In particular, stopped-flow assays revealed that trehalose slows down the binding of β-NADH to rbLDH, as well as its dissociation from the cofactor-enzyme complex. Conversely, the presence of the disaccharide does not alter the rate constant of the association to rbLDH of the substrate analogue oxamate. Furthermore, according to steady-state and stopped-flow assays, we present evidence that the increased velocity of oxaloacetate reduction triggered by trehalose is related to an improved occupancy by the substrate of the enzyme subunits, and by a favorable reciprocal orientation of β-NADH and oxaloacetate.