Mycobacterium Tuberculosis Protein Tyrosine Phosphatase Research Articles

Tyrosine inhibits the Mycobacterium tuberculosis protein tyrosine phosphatase MptpA

Tyrosine inhibits the Mycobacterium tuberculosis protein tyrosine phosphatase MptpA

Design, synthesis and biological evaluation of 3-substituted-2-thioxothiazolidin-4-one (rhodanine) derivatives as antitubercular agents against Mycobacterium tuberculosis protein tyrosine phosphatase B

Mycobacterium tuberculosis infections still pose a serious threat to human health. Combination therapies are effective medical solutions to the problem. Mycobacterium tuberculosis is an intracellular pathogen that mainly depends on a virulence factor (Mycobacterium tuberculosis protein tyrosine phosphatase B, MptpB) for its survival in the host. Therefore, MptpB inhibitors are potential components of tuberculosis combination treatments. Herein, a new series of MptpB inhibitors bearing a rhodanine group were developed using a structure-based strategy based on the virtual screening hit. The new MptpB inhibitors displayed potent MptpB inhibitory activities and great improvements in cell membrane permeability. The optimal compounds reduced the bacterial burden in a dose-dependent manner in a macrophage infection model, especially, a combination of compound 20 and rifampicin led to a bacterial burden reduction of more than 95%, greater than the reductions achieved with compound 20 or rifampicin alone. This research provides new insights into the rational design of new MptpB inhibitors and verifies that the MptpB inhibitor has a promising potential as a component of tuberculosis treatment.

The Mycobacterium tuberculosis protein tyrosine phosphatase MptpA features a pH dependent activity overlapping the bacterium sensitivity to acidic conditions

The Mycobacterium tuberculosis low-molecular weight protein tyrosine phosphatase (MptpA) is responsible for the inhibition of phagosome-lysosome fusion and is essential for the bacterium pathogenicity. This inhibition implies that M. tuberculosis is not exposed to a strongly acidic environment in vivo, enabling successful propagation in host cells. Remarkably, MptpA has been previously structurally and functionally investigated, with special emphasis devoted to the enzyme properties at pH 8.0. Considering that the virulence of M. tuberculosis is strictly dependent on the avoidance of acidic conditions in vivo, we analysed the pH-dependence of the structural and catalytic properties of MptpA. Here we show that this enzyme undergoes pronounced conformational rearrangements when exposed to acidic pH conditions, inducing a severe decrease of the enzymatic catalytic efficiency at the expense of phosphotyrosine (pTyr). In particular, a mild decrease of pH from 6.5 to 6.0 triggers a significant increase of K0.5 of MptpA for phosphotyrosine, the phosphate group of which we determined to feature a pKa2 equal to 5.7. Surface plasmon resonance experiments confirmed that MptpA binds poorly to pTyr at pH values < 6.5. Notably, the effectiveness of the MptpA competitive inhibitor L335-M34 at pH 6 does largely outperform the inhibition exerted at neutral or alkaline pH values. Overall, our observations indicate a pronounced sensitivity of MptpA to acidic pH conditions, and suggest the search for competitive inhibitors bearing a negatively charged group featuring pKa values lower than that of the substrate phosphate group.

An Insight into MptpB Inhibitors as a Key Strategy to Treat MDR and XDRTuberculosis.

Tuberculosis (TB) is a chronic, air-borne infectious disease caused by Mycobacterium tuberculosis (Mtb), which prominently affects the lungs and usually manifests in other organs. TB is preventable and curable but what makes it challenging is the emergence of resistance to the available treatment options. MDR-continued TB's expansion is one of the world's most pressing and difficult problems. Mtb revives via the reciprocity between Mycobacterium and host signalling pathways. Mtb secretes a virulence component called Mycobacterium tuberculosis protein tyrosine phosphatase (MptpB), which helps to survive against host macrophages. It indicates that targeting secreted virulence factors offers more benefits to circumvent the emergence of resistance. Many effective inhibitors of MptpA and MptpB have been discovered, providing a solid foundation for future research and development. Aside from possessing a structurally unique binding site in the Mtb enzyme, MptpB's minimal resemblance to other human phosphatases provides a broad platform for improving selectivity over host PTPs. We believe that addressing several parts of infection processes in the host and bacteria with combination therapy is the greatest way to reduce treatment burden and medication resistance. We have discussed the recent potent, selective, and efficacious MptpB inhibitors, such as natural and marine-based, isoxazole- linked carboxylic acid-based, oxamic acid-based, and lactone-based inhibitors, as potential strategies for treating TB.

1H-1,2,3-Triazole-4H-chromene-D-glucose hybrid compounds: Synthesis and inhibitory activity against Mycobacterium tuberculosis protein tyrosine phosphatase B.

A series of 1H-1,2,3-triazole-4H-chromene-D-glucose hybrid compounds 7a-w were synthesized using click chemistry of 2-amino-7-propargyloxy-4H-chromene-3-carbonitriles 5a-w. CuNPs@montmorillonite was used as a catalyst in the presence of DIPEA as an additive for this chemistry. All synthesized 1H-1,2,3-triazoles were examined for in vitro inhibition against Mycobacterium tuberculosis protein tyrosine phosphatase B (MtbPtpB). Nine 1H-1,2,3-triazoles, including 7c-e, 7h, 7i, and 7r-t, displayed remarkable inhibitory activity against MtbPtpB with IC50 < 10 μM; compound 7t exhibited the most potent inhibition in vitro with an IC50 value of 0.61 μM. Kinetic studies of the three most active compounds, 7c,h,t, showed their competitive inhibition toward the MtbPtpB enzyme. Induced-fit docking and MM-GBSA studies on the enzyme (PDB: 2OZ5) revealed that the most active compound 7t was more effective against MtbPtpB. Residues Arg64, Arg136, Ash165, Arg166, and Arg63 in the binding pocket were identified as potential ligand-binding hot-spot residues for ligand 7t. The binding free energy calculation by the MM-GBSA approach for ligand 7t indicated that Coulomb, lipophilic, and van der Waals energy terms are major contributors to the inhibitor binding. Furthermore, the stability of the ligand-protein complex and the structural insights into the mode of binding were confirmed by 300-ns molecular dynamics simulation of 7t/2OZ5.

Discovery of biphenyls bearing thiobarbiturate fragment by structure-based strategy as Mycobacterium tuberculosis protein tyrosine phosphatase B inhibitors

Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) is an important virulence factor that blocks the host immune response and facilitates M. tuberculosis growth in host cells. MptpB inhibitors are potential components of tuberculosis combination treatment. Herein, we present the development of new biphenyls MptpB inhibitors with greatly improved MptpB inhibition based on our reported thiobarbiturate lead 6 by rational design with the structure-based strategy. The eight biphenyls bearing thiobarbiturate fragment target compounds showed more potent MptpB inhibition (IC50: 1.18–14.13 μM) than the lead compound 6. Further molecular docking studies showed that compounds 13, 26, 27 and 28 had multiple interactions with active sites. Among them, compound 13 exhibited dose-dependent increased antituberculosis activity in mouse macrophages. The results displayed that the strategy of modification utilizing biphenyl scaffold was efficient. Our study identifies biphenyls bearing thiobarbiturate fragment as new MptpB inhibitors and verifies the therapeutic potential of antimycobacterial agent targeting MptpB.

A new anthraquinone from mangrove endophytic fungus Aspergillus sp. 16-5C

A new anthraquinone, asperquinone A (1), and four known anthraquinone derivatives 2–5 were isolated from the mangrove endophytic fungus Aspergillus sp. 16-5C. These structures were elucidated by spectroscopic analysis and the absolute configuration of 1 was unambiguously determined by ECD calculation. Compounds 1–5 showed no significant inhibitory effect against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB).

Identification of fusarielin M as a novel inhibitor of Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB)

The secreted Mycobacterium tuberculosis (Mtb) protein tyrosine phosphatase B (MptpB) is an essential virulence factor required for the intracellular survival of Mtb within host macrophages. MptpB has become a promising target for the development of novel anti-tuberculosis (TB) drugs. In this study, two new fusarielins, fusarielins M (1) and N (2), and a biogenetically related known compound, fusarielin G (3) were isolated from the marine-derived fungus Fusarium graminearum SYSU-MS5127. Their inhibitory effects on MptpB were evaluated. Among these compounds, fusarielin M substantially inhibited MptpB with a half-maximal inhibitory concentration (IC50) of 1.05 ± 0.08 μM, and an inhibition constant (Ki) of 1.03 ± 0.39 μM. Surface plasmon resonance analysis was used to characterize the interaction between fusarielin M and MptpB in vitro. Fusarielin M also exhibited cellular activity in blocking MptpB-mediated Erk1/2 and p38 inactivation in macrophages. Importantly, fusarielin M (20 μM) substantially reduced intracellular mycobacterial growth within macrophages, causing a 62% reduction in the bacterial burden. The binding mode of fusarielin M was further explored via molecular docking which suggested that fusarielin M binds to the active site of MptpB, forming a hydrogen bond with the side chain of Asp165; this is unique in the P-loop of MptpB compared to conventional human PTPs. The contact between fusarielin M and Asp165 in the catalytic loop provides a potential basis for inhibitor selectivity. Therefore, fusarielin M shows great potential as an anti-TB drug candidate.

Cis-2 and trans-2-eisocenoic fatty acids are novel inhibitors for Mycobacterium tuberculosis Protein tyrosine phosphatase A.

Small protein tyrosine phosphatase (PtpA) of Mycobacterium tuberculosis is attributed to the development of latent tuberculosis infection, and hence bocomes an interesting target for drug development. In this communication, inhibition of PtpA by naturally occurring fatty acids cis-2 and trans-2-eicosenoic acid is investigated. Mtb PtpA was heterologously expressed in Escherichia coli, and the activity of PtpA was inhibited by cis-2 and trans-2 eicosenoic fatty acids. Both compunds showed strong inhibition of PtpA activity with IC50 at low micromolar concentration. As comparison, trans-11-eicosenoic acid only slightly inhibit PtpA. In silico analysis confirmed the inhibition of PtpB by cis-2-eicosenoic acid by formation of several hydrogen bonds. These findings show that cis-2 and trans-2 eicosenoic fatty acids are potential candidates for latent tuberculosis inhibitors.

Polypropionate Derivatives with Mycobacterium tuberculosis Protein Tyrosine Phosphatase B Inhibitory Activities from the Deep-Sea-Derived Fungus Aspergillus fischeri FS452.

Fiscpropionates A-F (1-6), six new polypropionate derivatives featuring an unusual long hydrophobic chain, were isolated from the deep-sea-derived fungus Aspergillus fischeri FS452. Their structures were elucidated on the basis of spectroscopic analysis, and the absolute configurations were determined by J-HMBC analysis, electronic circular dichroism (ECD) calculations, and the modified Mosher's method. This is the first discovery of polypropionates from marine-derived fungi, and compounds 4 and 5 represent the first examples of polypropionate derivatives containing a 3-hydroxypiperidin-2-one as part of an imide linkage. In addition, compounds 1-4 exhibited significant inhibitory activities against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with the IC50 values of 5.1, 12, 4.0, and 11 μM, respectively. Enzyme kinetic experiments suggested that they all acted through a noncompetitive mechanism. A preliminary structure-activity relationship is discussed.

ATP synthase, an essential enzyme in growth and multiplication is modulated by protein tyrosine phosphatase in Mycobacterium tuberculosis H37Ra

Mycobacterium tuberculosis (Mtb) protein tyrosine phosphatase (PtpA) has so far been known to control intracellular survival of mycobacteria; whereas the ATP synthase which is essential for mycobacterial growth has recently been contemplated in developing a breakthrough anti-TB drug, diarylquinoline. Since both of these enzymes have been established as validated drug targets; we report a robust and functional relationship between these two enzymes through a series of experiments using Mtb H37Ra. In the present study we report that the mycobacterial ATP synthase alpha subunit is regulated by PtpA. We generated gene knock-out for the enzyme PtpA and subjected to determine the mycobacterial replication and the proteome profile of wild type, mutant (ΔptpA) and complemented (ΔptpA:ptpA) strains of Mtb H37Ra. A substantial amount of decrease in the protein level of ATP synthase alpha subunit (AtpA) in case of mutant H37Ra was observed, while the levels of the enzyme were either increased or remained unchanged, in wild type and in the complemented strains.

Ophiobolin-Type Sesterterpenoids from the Mangrove Endophytic Fungus Aspergillus sp. ZJ-68.

Eleven new ophiobolin-type sesterterpenoids, asperophiobolins A-K (1-11), along with 12 known analogues (12-23) were isolated from the cultures of the mangrove endophytic fungus Aspergillus sp. ZJ-68. The structures of the new compounds were elucidated through spectroscopic analyses, and their absolute configurations were assigned by a combination of Mo2(AcO)4-induced electronic circular dichroism spectra and quantum chemical calculations. Asperophiobolins A-D (1-4) represent the first examples possessing a five-membered lactam unit between C-5 and C-21 in ophiobolin derivatives. In the bioactivity assays, compounds 8-10 and 14-17 exhibited inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW 264.7 macrophage cells with IC50 values ranging from 9.6 to 25 μM, and compound 8 was found to show comparable inhibition of Mycobacterium tuberculosis protein tyrosine phosphatase B with an IC50 value of 19 μM.

Peptides and polyketides isolated from the marine sponge-derived fungus Aspergillus terreus SCSIO 41008

Two new isomeric modified tripeptides, aspergillamides C and D (compounds 1 and 2), together with fifteen known compounds (compounds 3–17), were obtained from the marine sponge-derived fungus Aspergillus terreus SCSIO 41008. The structures of the new compounds, including absolute configurations, were determined by extensive analyses of spectroscopic data (NMR, MS, UV, and IR) and comparisons between the calculated and experimental electronic circular dichroism (ECD) spectra. Butyrolactone I (compound 11) exhibited strong inhibitory effects against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with the IC50 being 5.11 ± 0.53 μmol·L−1, and acted as a noncompetitive inhibitor based on kinetic analysis.

Docking- and pharmacophore-based virtual screening for the identification of novel Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) inhibitor with a thiobarbiturate scaffold

Mycobacterium tuberculosis (Mtb) protein tyrosine phosphatase B (MptpB) is an important virulence factor for Mtb that contributes to survival of the bacteria in macrophages. The absence of a human ortholog makes MptpB an attractive target for new therapeutics to treat tuberculosis. MptpB inhibitors could be an effective treatment to overcome emerging TB drug resistance. Adopting a structure-based virtual screening strategy, we successfully identified thiobarbiturate-based drug-like MptpB inhibitor 15 with an IC50 of 22.4 μM, and as a non-competitive inhibitor with a Ki of 24.7 μM. Importantly, not only did it exhibit moderate cell membrane permeability, compound 15 also displayed potent inhibition of intracellular TB growth in the macrophage, making it an excellent lead compound for anti-TB drug discovery. To the best of our knowledge, this novel thiobarbiturate is the first class of MptpB inhibitor reported so far that leveraged docking- and pharmacophore-based virtual screening approaches. The results of preliminary structure-activity relationship demonstrated that compound 15 identified herein was not a singleton and may inspire the design of novel selective and drug-like MptpB inhibitors.

Synthesis, biological evaluation and molecular docking study of 1,2,3-1H-triazoles having 4H-pyrano[2,3-d]pyrimidine as potential Mycobacterium tuberculosis protein tyrosine phosphatase B inhibitors

Some heterocycles, namely 2-amino-4H-pyran-3-carbonitriles, were synthesized in a three-component reaction from substituted benzaldehydes, malononitrile, and ethyl acetoacetate. These heterocycles have been converted subsequently into 4H-pyrano[2,3-d]pyrimidine ring by ring-closing reaction with acetic anhydride in the presence of the concentrated sulfuric acid as catalyst. The successive alkylation reaction of lactam NH bond on pyrimidine-4-one ring was carried out using propargylic bromide in dry acetone in the presence of anhydrous potassium carbonate. The click chemistry of 3-propargyl-4H-pyrano[2,3-d]pyrimidine compounds has been accomplished by reaction with tetra-O-acetyl-α-d-glucopyranosyl azide using the metal-organic framework Cu@MOF-5 as a catalyst in absolute ethanol. All the synthesized 1H-1,2,3-triazoles 8a–y were screened for their in vitro Mycobacterium tuberculosis protein tyrosine phosphatase B (MtbPtpB) inhibition. Kinetic studies of the most active compounds 8v, 8x, and 8y showed their competitive inhibition toward the MtbPtpB enzyme. The detailed structure-activity relationship (SAR) in vitro and in silico studies suggested that the interaction of Arg63 amino acids with anion type of para-hydroxyl group via a salt bridge of iminium cation was essential for strong inhibitory activity against MtbPtpB.

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.

Two new bioactive steroids from a mangrove-derived fungus Aspergillus sp.

Two unusual naturally Diels-Alder additive steroids, ergosterdiacids A and B (1 and 2), constructing a 6/6/6/6/5 pentacyclic steroidal system, together with three known compounds (3–5) were obtained from the mangrove-derived fungus Aspergillus sp. Their structures were elucidated based on the comprehensive spectroscopic analysis, including 1D, 2D NMR and HRESIMS, as well as the quantum chemical ECD calculations. The plausible biosynthetic pathways of 1 and 2 were discussed. In the bioactivity assays, 1 and 2 exhibited potential in vitro inhibition activity against Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB) with an IC50 value of 15.1 and 30.1 μM, respectively. The inhibitory kinetic experiments indicated that both of them acted via a noncompetitive inhibition mechanism. Moreover, 1 and 2 showed strong in vitro anti-inflammatory effects by suppressing the NO production at 4.5 and 3.6 μM, respectively.

Peptides from the Soft Coral-associated Fungus Simplicillium sp. SCSIO41209

Five new peptides, Sinulariapeptides A−E together with seven known peptides (6–12) were isolated from the soft coral associated fungus Simplicillium sp. SCSIO 41209. The structures of the new compounds and their absolute configurations were established on the basis of spectroscopic analysis including NMR, MS and ECD. All the Compounds (except sinulariapeptides B−D) were tested for the inhibitory activities of Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB), and antifungal activities against five plant pathogenic fungi. Simplicilliumtides B and cyclo(L-Val-L-Pro) showed inhibitory activity with the IC50 values of 35.0 and 25.9 μM, sinulariapeptides A, simplicilliumtides J, verlamelins A and B exhibited potent inhibition against Colletotrichum asianum with the MIC values of range from 4.9 to 9.8 μg/mL and simplicilliumtides J, verlamelins A and B displayed inhibition against Pyricularia oryza Cav with the MIC values in the range of 19.5–78.1 μg/ml, respectively.

Peniisocoumarins A-J: Isocoumarins from Penicillium commune QQF-3, an Endophytic Fungus of the Mangrove Plant Kandelia candel.

Ten new isocoumarins, named peniisocoumarins A-J (1-9 and 11), along with three known analogues (10, 12, and 13) were obtained from the fermentation of an endophytic fungus, Penicillium commune QQF-3, which was isolated from a fresh fruit of the mangrove plant Kandelia candel. Their structures were elucidated through extensive spectroscopic analysis. The absolute configurations of 1-7 were determined by single-crystal X-ray diffraction and modified Mosher's method, and those of 8, 9, and 11 were assigned on the basis of experimental and calculated electronic circular dichroism data. Compounds 1 and 2 were unusual dimeric isocoumarins with a symmetric four-membered core. These isolated compounds (1-13) were evaluated for their cytotoxicity and enzyme inhibitory activities against α-glucosidase and Mycobacterium tuberculosis protein tyrosine phosphatase B (MptpB). Among them, compounds 3, 7, 9, and 11 exhibited potent inhibitory effects against α-glucosidase with IC50 values ranging from 38.1 to 78.1 μM, and compound 7 was found to inhibit MptpB with an IC50 value of 20.7 μM.

Synthesis of new C-dimethylated chalcones as potent antitubercular agents

A new class of C-dimethylated-chalcones (9a–q) were synthesized by using 2-hydroxy-3,5-dimethyl-4,6-dimethoxy acetophenone as a key intermediate. The compounds were screened for anti-tubercular activity against Mycobacterium tuberculosis strain (H37Rv) by Microplate Alamar Blue assay (MABA) method at a concentration of 100–0.8 µg/mL. The chalcones, 9a, 9b, 9c, 9k, 9o, and 9p were found to have higher antitubercular activity than the standard drugs, while the remaining compounds showed moderate activity. The antitubercular activity of the chalcones, 9b (MIC90 = 3.98 µM) and 9o (MIC90 = 3.84 µM) was found to be more than two-fold more active than the standard drugs, streptomycin (MIC90 = 10.75 µM) and ciprofloxacin (MIC90 = 9.43 µM), while their antitubercular activity was found to be more than six-fold more active than pyrazinamide (MIC90 = 25.38 µM). Further, the molecular docking studies employing Mycobacterium tuberculosis protein tyrosine phosphatase (MtbPtp) was carried out to observe docking scores.