Figure 10.

Nanobody effect on the catalytical function. (a) SsZntA was incubated with each Nb at a 1:2 molar ratio (i.e., 1.6 µM) and activity was assessed. Nb9 displayed an inhibitory effect of about 50% compared to untreated (no Nb) SsZntA. The difference between no Nb and Nb9 has a p value of <0.0001 (*), as assessed by a Dunn’s multiple comparison test. The ATPase inhibitor aluminum fluoride (AlF₃) is shown as a positive control, at 1.6 µM (corresponding to the concentration of the tested Nbs) and at 2 mM. (b) The inhibition stoichiometry was assessed as titration of SsZntA with different molar ratios of Nb9. The inhibitory effect appears when Nb9 is present at a molar ratio of 1:1 (ZntA:Nb9), and reaches the inhibitory plateau at a molar ratio of 1:3 (ZntA:Nb9), with an IC₅₀ = 1.57 molar ratio.