pH-dependent effects of the alkali-metal ions on intestinal brush-border sucrase.

The alkali-metal ions, Li+, Na+ and K+, exhibit complex pH-dependent effects on sucrase that seem to involve at least three separate types of mechanism and, perhaps, an equal number of specific metal-binding sites. In metal-free buffers, the alkali metals give mixed activating effects below pH 7 that involve: 1) Affinitytype (K-type) activation kinetics, where the apparent K,,, decreases as the metal concentration increases, and 2) Capacity-type (V-type) activation kinetics, where the apparent V,.= increases with the metal concentration. The K-type component disappears as the pH is raised from 5 to 7, suggesting the existence of a correlation between affinity-type metal activation and the deprotonation of an acidic group in the enzyme. On the contrary, the V-type component appears to be pH-independent and responsible for the essentially pure capacity-type kinetics observed at neutral pH. In lithium buffers at pH 5.2, kinetics are of the K-type only, strongly indicating that, in the presence of Li+, the V-component is fully saturated when the K-component is not. Therefore, the mixed-type kinetics seen below pH 7 may involve at least two distinct metalbinding activator sites. Above pH 7, all alkali-metal ions, particularly Li’, inhibit sucrase. Since these inhibitory effects take place at higher concentrations than those required to produce activation, velocity versus cation concentration curves are biphasic. In addition, some organic cations such as tert-butylammonium can interfere with the inhibitory action of the alkali metals above pH 7, further revealing the complexity of the interaction of monovalent cations with sucrase. The difficulty of finding truly inert buffers for the study of rabbit brush-border sucrase is discussed.