Natural Metabolites from Tabebuia aurea Reveal Isoform-Specific and Cooperative Modulation of Cathepsins B and L.

Authors

Marli de Fátima Correa Emiliano, Mariele Rondon Santos Gonçalves, Leice Milla Ribeiro de Novais, Marcos José Jacinto, Paulo Teixeira de Sousa, Mário Geraldo de Carvalho, Rayssa de Mello Lopes, Maurício Ferreira Marcondes Machado, Ana Laura Ferreira Vicco, Maria Luiza Gonçalves Gallego Rosa, Tiago Rodrigues, Renan Lira de Farias, André Silva Pimentel, Rayla Kelly Magalhães Costa, Virginia Claudia Paulino Silva, Wagner Alves de Souza Judice

Published in

Archives of biochemistry and biophysics. Pages 110971. Aug 14, 2026. Epub Aug 14, 2026.

Abstract

The development of isoform-selective inhibitors for homologous cysteine proteases remains challenging due to structural similarity and overlapping specificities. Metabolites isolated from Tabebuia aurea were investigated as modulators of Cathepsins B and L. Bioassay-guided fractionation yielded three compounds: 2,3-DH-5-HMB (1), veratric acid (2), and iridoid specioside (3). Enzymatic assays identified veratric acid as the most active compound (IC∼50∼ = 4.23 ± 0.17 μM for Cathepsin B and 1.28 ± 0.03 μM for Cathepsin L). Steady-state kinetic analyses revealed marked mechanistic divergence between isoforms. Against Cathepsin B, compounds 1 and 2 acted as competitive inhibitors with strong positive cooperativity, quantitatively captured by the cooperativity factor β (0.00041 and 0.0129, respectively), corresponding to 2,400-fold and 78-fold increases in affinity upon binding of a second inhibitor molecule. For compound 2, where extreme cooperativity precluded independent estimation of microscopic parameters, the quadratic cooperativity parameter γ (0.273 μM^-2) enabled robust quantification of the cooperative effect. In contrast, all three compounds inhibited Cathepsin L via a simple linear non-competitive mechanism (α ≈ 1), reflecting its more accessible active-site architecture. Cellular assays confirmed selective antiproliferative activity of veratric acid in K562 leukemia cells (EC∼50∼ = 12.47 μM) while sparing non-tumor HaCaT cells. Molecular docking and molecular dynamics simulations of 1:1 complexes provided structural context for the observed isoform-specific binding modes. These results demonstrate that structurally simple phenolic compounds can induce fundamentally different inhibition mechanisms in closely related enzymes, establishing a structure-mechanism framework for isoform-selective modulation of cysteine proteases.

PMID:
42600823
Bibliographic data and abstract were imported from PubMed on 15 Aug 2026.

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