作者: Svensson, Mattias ; Novello, Monica ; Bergonzini, Anna ; Norrby-Teglund, Anna ; Kaland, Lara ; Nilsson, Sara ; Engstrand, Olivia ; Tanaka, Emi ; Seidner, Lisa ; Palma Medina, Laura M.

Thrombomodulin (TM) is a membrane protein with significant roles in coagulation hemostasis and immune response. Its soluble form (sTM) has recently emerged as a key biomarker for severe invasive bacterial infections, including Necrotizing Soft Tissue Infections (NSTI). While various mechanical, chemical, and enzymatic mechanisms have been linked to TM shedding, this study investigates the direct impact of bacterial stimuli on soft tissue cells as primary sources of TM release. We stimulated organotypic models, composed of fibroblast and endothelial cells, with NSTI clinical isolates and found that while Group A Streptococcus and Escherichia coli had minimal effect on TM release, Staphylococcus aureus infection triggered a significant increase of sTM levels. We further assessed whether the secreted proteins of S. aureus led to higher TM levels by increased expression, increased cell toxicity, or direct cleavage of TM from the endothelial cell membrane. To investigate these mechanisms, we performed in vitro stimulations of endothelial monolayers with secreted proteins of two S. aureus isolates differing in their agr-system functionality. Our results indicate that S. aureus agr-regulated proteins induce TM shedding by direct cleavage from the cell membrane, an effect that was inhibited by metalloproteinase inhibitors. Stimulation with the pore-forming protein α-toxin showed similar results, suggesting a potential involvement of ADAM10 in TM cleavage. Additionally, we observed that other agr-regulated proteins can cleave TM directly. Altogether, this study reveals a pathogen-specific mechanism for TM release during S. aureus invasive infection, contributing to its elevated plasma levels and providing deeper insights into the pathophysiology of NSTI.

2026-02-02·INORGANIC CHEMISTRY

Substrate-Mimicking Peptides as MMP-1 Inhibitors: Impact of Zinc-Binding Group Position on Ternary Complex Stability

Article

作者: Potocki, Sławomir ; Gumienna-Kontecka, Elżbieta ; Potok, Paulina ; Woźniak-Laszczyńska, Wiktoria ; Palacios, Òscar ; Capdevila, Merce ; Wieczorek, Robert

Cancer remains a leading cause of global mortality, with metastasis accounting for nearly 90% of related deaths. Matrix metalloproteinases (MMPs), and in particular MMP-1, play a pivotal role in tumor progression by degrading extracellular matrix components through a Zn(II)-dependent catalytic mechanism. Targeting the Zn(II) ion in the active site represents a potential approach for inhibitor design. In this study, we designed and investigated substrate-mimicking peptide inhibitors incorporating cysteine residues as zinc-binding groups (ZBGs) at distinct positions: CPQGLRG (Inh4, P4), PQGLCGR (Inh2', P2'), and PQGLRGC (Inh4', P4'). Using different techniques (potentiometry, mass spectrometry, NMR spectroscopy, and density functional theory calculations), we evaluated binary and ternary complexes formed between these peptides, Zn(II), and an MMP-1 active-site model. All inhibitors formed monomeric and bis(ligand) binary Zn(II)-complexes, with Inh4 demonstrating the highest thermodynamic stability. In ternary systems, the MMP-1 active site model served as the primary Zn(II) ligand coordinating through three histidine residues and reproducing the binding mode of the native enzyme. The inhibitors bound in the secondary step as the fourth coordination site, displacing the catalytic water. Ternary complexes of all inhibitors were predominant species formed above pH 6, coinciding with the optimal pH for MMP-1's activity. Among the peptides, Inh4, which stabilized ternary complexes most effectively, coordinates Zn(II) via its N-terminal amine. This binding mode is analogous to the strategy of tissue inhibitors of metalloproteinases. In contrast, Inh2' and Inh4' required structural rearrangements for Zn(II) coordination and formed less stable complexes due to steric constraints. The findings of this study identify N-terminal cysteine as the most effective ZBG placement for stabilizing Zn(II)-MMP-1 complexes, highlighting Inh4 as a promising lead for peptide-based MMP-1 inhibition. This work provides preliminary insights to guide the rational design of selective metalloproteinase inhibitors with therapeutic potential in cancer treatment.

2024-04-01·CURRENT OPINION IN BIOTECHNOLOGY

Engineering metalloproteinase inhibitors: tissue inhibitors of metalloproteinases or antibodies, that is the question

Review

作者: Hilpert, Gregory A ; Raeeszadeh-Sarmazdeh, Maryam ; Mosca, Ethan R ; Kalantar, Masoud

Targeting metalloproteinases (MPs) has been the center of attention for developing therapeutics due to their contribution to a wide range of diseases, including cancer, cardiovascular, neurodegenerative disease, and preterm labor. Protein-based MP inhibitors offer higher stability and selectivity, which is critical for developing efficient therapeutics with low off-target effects. Tissue inhibitors of metalloproteinases (TIMPs), natural inhibitors of MPs, and antibodies provide excellent protein scaffolds for engineering selective or multispecific MP inhibitors. Advances in protein engineering and design techniques, such as rational design and directed evolution using yeast display to develop potent MP inhibitors, are discussed, including but not limited to loop grafting, swapping, and counterselective selection.

100 项与 01-07A 相关的药物交易

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