Synthetic thiosemicarbazones as a new class of Mycobacterium tuberculosis protein tyrosine phosphatase A inhibitors.

Mycobacterium tuberculosis secretes two protein tyrosine phosphatases as virulence factors, PtpA and PtpB. Inhibition studies of these enzymes have shown significant attenuation of the M. tuberculosis growth in vivo. As PtpA mediates many effects on the regulation of host signaling ensuring the intracellular survival of the bacterium we report, for the first time, thiosemicarbazones as potential novel class of PtpA inhibitors. Several compounds were synthesized and biologically evaluated, revealing interesting results. Enzyme kinetic assays showed that compounds 5, 9 and 18 are non-competitive inhibitors of PtpA, with Ki values ranging from 1.2 to 5.6 µM. Modeling studies clarified the structure-activity relationships observed in vitro and indicated a possible allosteric binding site in PtpA structure. To the best of our knowledge, this is the first disclosure of potent non-competitive inhibitors of PtpA with great potential for future studies and development of analogues.

S-nitrosylation of Mycobacterium tuberculosis tyrosine phosphatase A (PtpA) induces its structural instability.

S-nitrosylation is associated with signal transduction and microbicidal activity of nitric oxide (NO). We have recently described the S-nitrosylation of Mycobacterium tuberculosis protein tyrosine phosphatase A, PtpA, an enzyme that plays an important role in mycobacteria survival inside macrophages. This post-translational modification decreases the activity of the enzyme upon modification of a single Cys residue, C53. The aim of the present work was the investigation of the effect of S-nitrosylation in PtpA kinetic parameters, thermal stability and structure. It was observed that the K(M) of nitrosylated PtpA was similar to its unmodified form, but the V(max) was significantly reduced. In contrast, treatment of PtpA C53A with GSNO, did not alter either K(M) or V(max). These results confirmed that PtpA S-nitrosylation occurs specifically in the non-catalytic C53 and that this modification does not affect substrate affinity. Using circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy techniques it was shown that PtpA S-nitrosylation decreased protein thermal stability and promoted a local effect in the surroundings of the C53 residue, which interfered in both protein stability and function.

Antiradical effect and phytochemical characterisation of the leaves of Parkia platycephala Benth.

The objective of this study was to isolate and identify compounds present in the leaves of Parkia platycephala Benth (Leguminosae-Mimosoideae) and evaluate the antiradical potential of the derived extracts and fractions. Seven compounds were successfully isolated from the ethanolic leaf extract, comprising gallic acid, flavonoids, triterpenoids, and phytosteroids. Structural identification was accomplished through comprehensive spectroscopic data analysis, including NMR and mass spectrometry (ESI-MS), and comparison with existing literature. The antiradical efficacy of both the extract and its fractions was determined in vitro through assays measuring 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and quantifying total phenolics via UV/Vis, revealing notable antioxidant activity in extracts from P. platycephala Benth, especially in the EtOH, EtOAc, and aqueous fractions, attributed to their high phenolic content and prevalence of flavonoids. These findings contribute to a deeper understanding of Parkia's chemical composition and its potential biological benefits.

Targeting protein tyrosine phosphatases for the development of antivirulence agents: Yersinia spp. and Mycobacterium tuberculosis as prototypes.

Protein phosphorylation mediated by protein kinases and phosphatases has a central regulatory function in many cellular processes in eukaryotes and prokaryotes. As a result, several diseases caused by imbalance in phosphorylation levels are known, especially due to protein tyrosine phosphatases (PTPs) activity, an important family of signaling enzymes. Furthermore, over the last decades several studies have shown the main role of PTPs in pathogenic bacteria: they are associated with growth, cell division, cell wall biosynthesis, biofilm formation, metabolic processes, as well as virulence factor. In this way, PTPs have ascended as targets for antibacterial drug design, particularly in view of the antibiotic resistance in pathogenic bacteria, which demands novel therapeutics strategies. Targeting secreted PTPs is an antivirulence strategy to combat the emergence of antimicrobial resistance (AMR). This review focuses on the recent advances in understanding the role of PTPs and the approaches to target them, with an emphasis in Yersinia spp. and Mycobacterium tuberculosis pathogenesis.

Discovery of Mycobacterium tuberculosis protein tyrosine phosphatase B (PtpB) inhibitors from natural products.

Protein tyrosine phosphatase B (PtpB) is one of the virulence factors secreted into the host cell by Mycobacterium tuberculosis. PtpB attenuates host immune defenses by interfering with signal transduction pathways in macrophages and, therefore, it is considered a promising target for the development of novel anti-tuberculosis drugs. Here we report the discovery of natural compound inhibitors of PtpB among an in house library of more than 800 natural substances by means of a multidisciplinary approach, mixing in silico screening with enzymatic and kinetics studies and MS assays. Six natural compounds proved to inhibit PtpB at low micromolar concentrations (< 30 µM) with Kuwanol E being the most potent with K i = 1.6 ± 0.1 µM. To the best of our knowledge, Kuwanol E is the most potent natural compound PtpB inhibitor reported so far, as well as it is the first non-peptidic PtpB inhibitor discovered from natural sources. Compounds herein identified may inspire the design of novel specific PtpB inhibitors.

Synthetic chalcones and sulfonamides as new classes of Yersinia enterocolitica YopH tyrosine phosphatase inhibitors.

YopH plays a relevant role in three pathogenic species of Yersinia. Due to its importance in the prevention of the inflammatory response of the host, this enzyme has become a valid target for the identification and development of new inhibitors. In this work, an in-house library of 283 synthetic compounds was assayed against recombinant YopH from Yersinia enterocolitica. From these, four chalcone derivatives and one sulfonamide were identified for the first time as competitive inhibitors of YopH with binding affinity in the low micromolar range. Molecular modeling investigations indicated that the new inhibitors showed similar binding modes, establishing polar and hydrophobic contacts with key residues of the YopH binding site.