ActiProbe: Activity-Based Probes as chemical tools for biomarker discovery in Chronic Obstructive Pulmonary Disease

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3-Oxo-β-Sultams and 4-Oxo-β-Lactams as chemical tools for activity-based protein profiling of serine hydrolases

Publication . Carvalho, Luís Miguel Afonso Ramos de; Moreira, Rui Ferreira Alves; Penque, Deborah; Lucas, Susana Dias

Activity-based protein profiling (ABPP) is a technique that analyzes the dynamics in enzymatic activity in complex proteomes by using small molecular probes, deemed activity-based probes (ABPs), containing a reactive group to covalently bind enzyme catalytic residues, a tag for detection of labeled targets and a linker as a spacer and also specificity-enhancing. Different reactive groups have been developed to engage a wide range of enzymatic families but there is a constant need to create new chemical tools to expand the pool of engageable targets. In this work we evaluated two 4-membered ring chemotypes as new reactive groups for ABPP of serine hydrolases. The 3-Oxo-β-Sultam was revealed to be a highly reactive chemotype which labels a wide range of proteins with limited target occupancy. A crystallographic analysis of the reaction of 3-Oxo-β-Sultams with elastase enzymes revealed a previously unknown mechanism of inhibition of these enzymes by sulfonylation, suggesting 3-Oxo-β-Sultam compounds could be used to expand the pool of available sulfonylating tools in chemical biology. 4-Oxo-β-Lactams were shown to potently hit a selected group of serine hydrolase with high target occupancy, including human neutrophil elastase (HNE) and members of the ABHD and DPP families of enzymes. A competitive-ABPP approach revealed high potency of a library of 4-Oxo-β-Lactams to target these enzymes. 4-Oxo-β-Lactams were identified as a new chemotype for DPP8 and DPP9 inhibition. Crystallography experiments revealed a new binding mode of these enzymes with 4-Oxo-β-Lactams, highlighting that these compounds could be used to pursue selective DPP8 or DPP9 inhibitors, a highly pursued field in current medicinal chemistry.

2020-02Doctoral thesis

Open access

3-Oxo-ß-sultam as a Sulfonylating Chemotype for Inhibition of Serine Hydrolases and Activity-Based Protein Profiling

Publication . Carvalho, Luís; Almeida, Vanessa Tavares; Brito, José A.; Lum, Kenneth; Oliveira, Tânia F.; Guedes, Rita C.; Gonçalves, Lídia; Lucas, Susana Dias; Cravatt, Benjamin; Archer, Margarida; Moreira, Rui

3-Oxo-β-sultams are four-membered ring ambident electrophiles that can react with nucleophiles either at the carbonyl carbon or at the sulfonyl sulfur atoms, and that have been reported to inhibit serine hydrolases via acylation of the active-site serine residue. We have developed a panel of 3-oxo-β-sultam inhibitors and show, through crystallographic data, that they are regioselective sulfonylating electrophiles, covalently binding to the catalytic serine of human and porcine elastases through the sulfur atom. Application of 3-oxo-β-sultam-derived activity-based probes in a human proteome revealed their potential to label disease-related serine hydrolases and proteasome subunits. Activity-based protein profiling applications of 3-oxo-β-sultams should open up new opportunities to investigate these classes of enzymes in complex proteomes and expand the toolbox of available sulfur-based covalent protein modifiers in chemical biology.

2020-03-16Journal article

Restricted access