Despite the very high degree of conservation among this specific family of nucleotidyltransferases, MZP harbors a 5-to 25-fold gain in specificity for HCE when compared to other GTases. This result additionally supports the presence of an allosteric MZP binding pocket. Oddly enough, the lone GTase domain of HCE is less susceptible by 10-fold to MZP inhibition than the full-length HCE. This evidence, not only supports the presence of an allosteric site, but can also provide additional information about its localization on HCE. It is tempting to speculate that MZP could bind near the N-terminal of the GTase domain or on a region of interaction between both the RTase and GTase domains since the abolition of the RTase domain reduces HCE MZP susceptibility. In order to gain additional details on the MZP main binding site, molecular docking could be used. Unfortunately, the structure of both the RTase and the GTase domain are separately available, but the structure of the full-length protein is still not available. Nevertheless, we ran a molecular docking experiment of MZP on the GTase domain from HCE. The lack of RTase domain, which is predicted to participate in MZP binding, and the moderate flexibility of the N-terminal domain, which introduces a structural incertitude, does not allow for definitive conclusions to be reached but it is interesting to note that, in preliminary experiments, MZP favorably docks on the N-terminal region of the HCE GTase. Together, our results reveal that MZP inhibits the HCE GTase activity with a 5-to 25-fold specificity in comparison to other GTases. Although more work is yet required to confirm our hypothesis, these results raise the possibility that the GTase inhibition could be mediated by a conformational change hindrance upon binding of MZP to an allosteric binding site that is speculated to reside near the RTase-GTase inter-domain. Nevertheless, mizoribine is one of the first compounds to demonstrate a certain degree of specificity toward a single GTase, despite the high degree of conservation of this crucial family of enzyme. MZP displays a higher in vitro inhibition potency for the GTase reaction in comparison to the complete RNA capping reaction. This may simply be due to our experimental BML-284 customer reviews conditions where the RTase activity of HCE, which is partially inhibited by free Mg2+, becomes the rate-limiting step. However, in cellulo the RTase harbors a higher turnover rate than the GTase, which catalyzes the limiting step in RNA capping. In a cellular context we expect the efficiency of MZP to be dictated solely by its interaction with the GTase. Historically, very few GTase inhibitors have been developed, neither as scientific tools nor as Nigericin (sodium salt) therapeutic agents.