| Full text | |
| Author(s): |
Chagas, Rafael S.
;
Marana, Sandro R.
Total Authors: 2
|
| Document type: | Journal article |
| Source: | PLoS One; v. 20, n. 3, p. 11-pg., 2025-03-25. |
| Abstract | |
Here we demonstrate that Tris (2-amino-2-(hydroxymethyl)-1,3-propanediol), largely used as a buffering agent, is a linear mixed inhibitor (Ki = 12 +/- 2 mM and alpha = 3 +/- 1) of the GH1 beta-glucosidase from the insect Spodoptera frugiperda (Sf beta gly). Such an inhibition mechanism implies the formation of a non-productive ESI complex involving Sf beta gly, substrate, and Tris. In addition, Tris binding reduces by 3 fold the enzyme affinity for the substrate. Hence, at concentrations higher than the Ki, Tris can completely abolish Sf beta gly activity, whereas even at lower concentrations the presence of Tris causes underestimation of beta-glucosidase kinetic parameters (Km and kcat). In agreement with the inhibition mechanism, computational docking showed that Tris could bind to a pocket placed at the lateral of the active site opening in the Sf beta gly-substrate complex, hence leading to the formation of an ESI complex. In agreement with the crystallographic data available, computational docking also showed that Tris may find binding spots in the interior of the active site of the Sf beta gly and several GH1 beta-glucosidases. Moreover, the variety of their active site shapes results in a multiplicity of binding profiles, foreseeing different inhibition mechanisms. Thus, Tris inhibition may affect other GH1 beta-glucosidases. This remark should be taken into account in their study, highlighting the importance of the appropriate buffer for accurate enzyme characterization. (AU) | |
| FAPESP's process: | 21/10577-0 - Biology of Bacteria and Bacteriophages Research Center |
| Grantee: | Shaker Chuck Farah |
| Support Opportunities: | Research Grants - Research, Innovation and Dissemination Centers - RIDC |
| FAPESP's process: | 21/03967-6 - Characterization of a beta-glycosidase dimer interface |
| Grantee: | Sandro Roberto Marana |
| Support Opportunities: | Regular Research Grants |