Recent studies have found that the suppression of phosphatase and tensin homolog is one of the most effective single-gene approaches for promoting optic nerve regeneration. This effect is primarily mediated through the activation of the protein kinase B/phosphoinositide 3-kinase/mammalian target of rapamycin signaling pathway. The purpose of this article is to elucidate how the downregulation of phosphatase and tensin homolog is involved in each key phase of optic nerve regeneration and to summarize the potential targets for therapeutic interventions in this process. Optic nerve regeneration progresses through five phases: stress response, growth navigation, nerve regeneration, synaptic reconstruction, and remyelination. During the stress response phase, the suppression of phosphatase and tensin homolog enhances the survival of retinal ganglion cells and promotes the proliferation of microglia. In the nerve regeneration phase, reduced levels of phosphatase and tensin homolog facilitate mitochondrial transport, while inhibition of the phosphatase and tensin homolog-L isoform specifically promotes mitophagy. During the synaptic reconstruction phase, the deletion of phosphatase and tensin homolog modulates the synthesis of axon extension-related proteins and stabilizes microglial microtubules, thereby accelerating the clearance of damaged synapses and the formation of new ones. During the remyelination phase, the knockout of phosphatase and tensin homolog promotes the proliferation of oligodendrocyte progenitor cells and the differentiation of oligodendrocytes, relieving myelination obstruction. This paper also discusses current strategies and translational challenges for neuron-specific inhibition of phosphatase and tensin homolog, including off-target effects, delivery precision, and long-term safety. By integrating molecular insights with emerging bioengineering approaches, this paper provides a framework for developing targeted therapies for optic nerve regeneration and broader applications in the field of central nervous system regeneration.

ABSTRACTRett syndrome (RTT) is a severe neurodevelopmental disorder in which current therapeutic strategies remain largely focused on providing symptomatic relief without addressing underlying disease mechanisms. In contrast, we have identified the protein tyrosine phosphatase PTP1B as a mechanism-based therapeutic target and evaluated a class of selective, allosteric small-molecule inhibitors in female murine models of RTT. We show that one of these compounds localizes to brain regions central to motor coordination and cardio-respiratory control, which are core domains of RTT pathology. Pharmacological inhibition of PTP1B produces robust and sustained improvement in multiple disease symptoms, including muscle weakness, motor and coordination deficits, and cardiac and respiratory dysfunction. Concordant results obtained with genetic ablation of PTP1B, with effects maintained for over one year, demonstrate that phenotypic rescue arises from on-target modulation of disease-relevant signaling. Mechanistically, PTP1B inhibition is known to normalize neurotrophic and metabolic pathways, including TRKB and insulin/leptin signaling, thereby restoring circuit-level function. These findings establish PTP1B as a clinically actionable, disease-modifying target and demonstrate that selective, allosteric inhibition of a protein tyrosine phosphatase can achieve durable therapeutic benefit in vivo. This work provides a strong rationale for the clinical evaluation of PTP1B inhibitors as a mechanism-based treatment strategy for RTT.One Sentence SummaryWe have validated inhibition of PTP1B as a mechanism-based therapeutic strategy that alleviates a wide range of symptoms in a mouse model of Rett syndrome.

Vascular endothelial cells form a selective barrier that regulates the passage of substances and leukocytes between the bloodstream and surrounding tissues, thereby maintaining vascular homeostasis. Although endothelial barrier dysfunction is implicated in numerous diseases, the molecular mechanisms that protect against such dysfunction remain incompletely defined. Thrombin, an inflammatory mediator, increases endothelial permeability by inducing myosin light chain (MLC) phosphorylation through Rho/Rho-associated kinase (Rho-kinase)-mediated inhibition of myosin phosphatase. This process disrupts vascular endothelial cadherin (VE-cadherin)-based junctions and promotes radial stress fiber formation. Here, we demonstrate that the green tea catechin (–)-epigallocatechin-3-gallate (EGCG) reduces phosphorylation of the myosin phosphatase regulatory subunit MYPT1 at inhibitory sites and suppresses Rho-kinase signaling in endothelial cells. Together, these EGCG-mediated effects reduce MLC phosphorylation, inhibit radial stress fiber formation, and preserve VE-cadherin-mediated cell–cell adhesion, thereby maintaining endothelial barrier integrity.

It is increasingly recognized that the 'omic analysis of whole blood has applications for precision medicine and disease phenotyping. Despite this realization, whole blood is generally viewed as a challenging analytical matrix in comparison to plasma or serum. Moreover, proteomic analyses of whole blood have almost exclusively focused on (non)targeted analyses of protein abundances and much less on post-translational modifications (PTMs). Here, we developed a streamlined workflow for processing 20 microliters of venous blood collected by volumetric absorptive microsampling that incorporates serial trypsinization and N-glycopeptide and phosphopeptide enrichment and avoids laborious sample dry-down or cleanup steps. As many as 10,000 analytes (reported as protein groups, glycopeptidoforms, and phosphosites) can be quantified by liquid chromatography-tandem mass spectrometry in under 2 h of MS acquisition time. Using these methods, we explored the stability of "dried" and "wet" blood proteomes, as well as the effects of ex vivo inflammatory stimulus or phosphatase inhibition. Multiomics factor analysis enabled facile identification of analytes that contributed to interindividual variability of the blood proteomes, including N-glycopeptides that distinguish immunoglobulin heavy constant alpha 2 allotypes. Collectively, our results help to establish feasibility and best practices for the integrated MS-based quantification of proteins and PTMs from dried blood.

Protein phosphorylation regulates diverse cellular processes, yet its analysis at the single-cell level remains challenging due to the low abundance of phosphoproteins. Here, we present a highly sensitive system for profiling phosphorylation of kinase substrates in individual cells. The method integrates fluorescence labeling of single-cell proteomes, immunoprecipitation using antibodies recognizing phosphorylation within specific amino acid motifs, miniaturized SDS-PAGE, and single-molecule detection using a custom-built light-sheet fluorescence microscope. We applied this approach to analyze substrates of casein kinase 2 (CK2) in HeLa cells treated with the phosphatase inhibitor calyculin A. Bulk and pseudo-single-cell analyses confirmed treatment-induced accumulation of phosphorylated CK2 substrates and demonstrated quantitative performance over biologically relevant input ranges. Importantly, true single-cell measurements revealed heterogeneous phosphorylation patterns across molecular weight regions, highlighting cell-to-cell variability in CK2 signaling that is obscured in bulk analyses. This platform enables profiling of the phosphorylation states of a wide range of kinase substrates in individual cells and provides a foundation for dissecting heterogeneous signaling dynamics.

Protein tyrosine phosphatases (PTPs) can be involved in the development of different human diseases, including cancer. In this study, in vitro experiments were performed to evaluate the inhibitory effect of the calix[4]arene α-ketophosphonic acids on the activities of PTP1B, TC-PTP, SHP2, and MEG2 under irradiation of 365nm UV light. Among the compounds tested, calix[4]arene bis-α-ketophosphonic acids substituted at the lower rim by two n-propyl or n-butyl groups significantly increased the UV-induced inhibition of the PTPs, demonstrating IC50 values in the low-micromolar range. The observed pseudo-first-order rate constants for the loss of activity of PTP1B and SHP2 depended linearly on the inhibitor concentration. The value of the second-order rate constant for SHP2 exceeded that for PTP1B. This work demonstrates that calix[4]arene can be used as a scaffold for designing photoactivatable inhibitors targeting PTPs.

Pancreatic ductal adenocarcinoma (PDAC) remains a therapeutic challenge, and the aggressive basal-like/mesenchymal subtype is particularly refractory to chemotherapy, underscoring the need for novel therapies. Leveraging genetic screens, we identified protein phosphatase 2A (PP2A) catalytic subunit PPP2CA as a target. Pharmacological PP2A inhibition selectively impaired the growth of mesenchymal PDAC cells. To delineate the mechanisms underlying sensitivity to the PP2A inhibitor LB100, we employed a dual-pronged strategy. Functional characterization revealed metabolic reprogramming coupled with endoplasmic reticulum (ER) stress and cell death induction. Genome-wide genetic screens identified key modifiers of LB100 sensitivity, implicating transcriptional regulators, mRNA processing, translation, and metabolism. Based on expression data linking PP2A to splicing and transcriptional regulation, we prioritized these processes for validation. Mesenchymal PDAC cells exhibited enhanced splicing following PP2A inhibition. Notably, we identified enhanced transcriptional elongation upon LB100 treatment, particularly of short genes, driven by cyclin-dependent kinase 9 (CDK9). Our findings support a reciprocal regulatory relationship between PP2A and CDK9 that connects to the activation of ER stress response factors, including activating transcription factor 4 (ATF4). These results establish PP2A as a druggable target in mesenchymal PDAC cells and reveal a role of LB100-induced transcriptional elongation and splicing, providing a mechanistic basis to guide future therapy development.

Drought can disrupt biogeochemical functioning in forest ecosystems by limiting microbial activity and extracellular enzyme production. We investigated the effects of simulated moisture deficit on the activity of five enzymes involved in C, N and P acquisition in a deadwood-soil system: β-glucosidase (BG), β-D-cellobiosidase (CB), β-xylosidase (XYL), N-acetyl-β-D-glucosaminidase (NAG) and phosphatase (PH). Deadwood of six temperate tree species (broadleaf and conifer) was exposed to drought and control conditions over two years (2023-2024), and enzyme activities were measured seasonally in both deadwood and the underlying soil. Drought conditions led to a pronounced reduction in the activity of all analyzed enzymes in deadwood, frequently exceeding 50% compared to control treatments, which indicates strong moisture limitation of microbial processes during wood decomposition. Enzymatic activity in soil beneath deadwood also decreased, although the smaller absolute changes observed in soil are likely related to lower initial activity levels rather than increased resistance to drought. The extent of decline varied among enzymes and wood species, with β-glucosidase and β-xylosidase showing differences of approximately 30-40% between treatments in soil, and with some species (e.g. beech and spruce) exhibiting weaker responses to drought. Enzyme activity in both substrates followed a consistent seasonal pattern, with maxima in summer and minima in winter. The increasing divergence between control and drought treatments in the second-year highlights cumulative effects of prolonged water limitation. The results demonstrate that extracellular enzyme activity is highly sensitive to moisture availability and that deadwood and underlying soil respond differently to drought. This provides mechanistic insight into how drought may alter microbial functioning and decomposition dynamics at the deadwood-soil interface in forest ecosystems. General Linear Models, correlation analysis and PCA consistently indicated moisture as a major factor associated with enzymatic variation, whereas temperature showed no significant relationships with enzyme activities. Reduced enzyme activity under drought suggests a limitation of microbial decomposition processes and nutrient acquisition, particularly for carbon-, nitrogen- and phosphorus-related pathways, potentially constraining microbial metabolism and slowing organic matter turnover at the deadwood-soil interface.

BackgroundThe nucleocapsid (N) protein of coronavirus harbors a conserved serine/arginine (SR)–rich motif whose phosphorylation by GSK-3 is essential for viral transcription and replication. Our previous studies in Severe acute respiratory syndrome coronavirus 1 (SARS-CoV-1) and the JHM strain of mouse hepatitis virus revealed a phosphorylation-to-dephosphorylation transition during the viral life cycle, with newly synthesized N highly phosphorylated and virion-associated N hypophosphorylated. Here, we characterize this transition in SARS-CoV-2 and define its functional significance.MethodsUtilizing high-resolution gel analysis and the phospho-specific antibody, the phosphorylation levels of SARS-CoV-2 N proteins in the virus-infected Calu-3 cells and secreted virions from culture supernatants were compared between different viral strains. This phosphorylation-to-dephosphorylation transition was also verified in the SARS-CoV-2 virus-like particle (SC2-VLP) platform expressing N, membrane (M), envelope, and spike proteins. In this VLP system, the substantial contribution of N dephosphorylation status to particle secretion, either by blocking GSK-3 activity or phospho-related mutants, was assessed. With different viral structural protein-expressing clones and phosphatase inhibitors, we characterized the key viral factor and host phosphatase to mediate the N dephosphorylation process. Furthermore, we evaluated the antiviral efficacy of phosphatase inhibitors using the virus-infected Calu-3 culture.ResultsOur results showed that the phosphorylation change of SR motif in N was observed across multiple strains and recapitulated in the SC2-VLP system. GSK-3 inhibition or a phospho-deficient N mutant, but not a phospho-mimetic one, enhanced VLP release, indicating that N dephosphorylation promotes virion secretion. Mechanistically, the M protein, via its C-terminal domain, interacts with phosphorylated N to recruit protein phosphatase 2A (PP2A) to the ERGIC, facilitating N dephosphorylation. Inhibition of PP2A, either by inhibitors or siRNA, impaired the M-induced N dephosphorylation, thereby suppressing viral assembly and progeny virion production. Blockade of PP2A also indirectly reduced N phosphorylation through the Akt-mediated inhibition of GSK-3, resulting in decreased genomic RNA synthesis.ConclusionsCollectively, these findings establish N dephosphorylation as a critical regulatory step in SARS-CoV-2 assembly and virion release. In ERGIC, this M protein-mediated process recruits host PP2A to dephosphorylate N, thus supporting PP2A as a promising pan-coronavirus antiviral target.Graphical Supplementary InformationThe online version contains supplementary material available at 10.1186/s12929-026-01255-w.

Hyperosmolarity caused by drought, high salinity, or cold stress inhibits plant growth and crop productivity. A conserved protein-kinase cascade of cytosolic B-RAFs and SnRK2s is rapidly activated upon osmotic stresses to initiate downstream adaptive responses, which represents one of the fastest known responses to osmotic stress in plants. How the kinase cascade is activated by osmotic stress is unknown. Here, we show that Arabidopsis B4 subgroup RAFs have intrinsically disordered regions and directly sense both ionic and nonionic hyperosmolarity by reversible condensation. B4-RAFs recruit and cocondense with subclass-I SnRK2s to phosphorylate and turn on SnRK2s, evading the noncondensable inhibitory A-clade PP2C phosphatases. This straightforward osmosensing and relaying module can be fully reconstituted in Escherichia coli by coexpressing three components or in solution in a test tube using recombinant proteins. Our findings identify B-RAFs as the chief cellular osmosensors that detect low water potential by cocondensation, forming a signal hub with SnRK2s to orchestrate adaptive responses in plants, and represent an evolutionarily conserved osmosensing mechanism across kingdoms.

The endangered Uncaria gambir (W.Hunter) Roxb. (U. gambir) remains largely unexplored in chemical constituents and bioactivities. Therefore, fifteen compounds (1–15), comprising monoterpene indole alkaloids, lignans, flavonoids, triterpenoids, coumarins, and fatty acids, were isolated from this species. Their inhibitory effects against cell division cycle 25B (CDC25B) phosphatase and vasodilatory activities were then evaluated. Results showed that compound 1 (strictosamide) exhibited a notable inhibitory effect on CDC25B phosphatase, with the IC50 value of 5.4 μM, comparable to the positive control (Na3VO4). Compound 3 (3α-dihydrocadambine) effectively revealed strong vasodilatory activity with EC50 values of 2.4 μM and 2.1 μM, respectively, similar levels comparable to the positive control (verapamil). Interestingly, these activities of compounds 1 and 3 were evaluated for the first time in this work. Moreover, network pharmacology and molecular docking predicted compound 1 to correlate with the p53-Hippo-TGF-β antitumor signalling pathway and compound 3 with the cAMP/cGMP-PKG cardiovascular signalling pathway. This work provides preliminary experimental data for the development of potential lead compounds from U. gambir.

Neuronal death triggered by hypoxia-ischemia (HI) is a major cause of neonatal mortality and long-term neurological deficits. Excessive autophagy plays a pathogenic role in neonatal hypoxic-ischemic encephalopathy (HIE), and inhibition of phosphatase and tensin homolog deleted on chromosome TEN (PTEN) nuclear translocation has been shown to suppress autophagy. Our recent study demonstrated that blocking PTEN nuclear import with the peptide Tat-K13 mitigates HI-induced behavioral impairments. However, the underlying mechanism remains unclear. Here, we found that HI activated the p-JUN-SESN2-AMPK signaling pathway in both in vivo and in vitro models of neonatal hypoxic-ischemic brain damage (HIBD). Downregulation of JUN reduced neuronal loss and improved behavioral outcomes in HIBD rats. Furthermore, Tat-K13-mediated blockade of PTEN nuclear translocation attenuated HI-induced activation of the p-JUN-SESN2-AMPK pathway and suppressed autophagy. Notably, the neuroprotective and behavioral benefits conferred by Tat-K13 were achieved through autophagy inhibition resulting from suppression of this signaling cascade. These findings identify targeting PTEN nuclear import with Tat-K13 as a potential therapeutic strategy for neonatal HIE, acting via the p-JUN-SESN2-AMPK-autophagy axis to promote neuronal survival and functional recovery.

The GID/CTLH E3 ligase complex is implicated in several biological processes, yet its full substrate repertoire remains poorly defined. We recently identified the complex as a broad modulator of macrophage responses to Mycobacterium tuberculosis (Mtb) infection. Here, we use label-free proteomics and diGly capture analysis of Mtb-infected macrophages to define the GID/CTLH-dependent ubiquitylome. We identify thousands of dynamically altered ubiquitylation sites, with strong enrichment among proteins involved in cellular metabolism and innate immune signaling. Concurrent proteome analysis revealed extensive rewiring in GID/CTLH-deficient macrophages, with >90% of enriched pathways among increased proteins consisting of metabolic targets. Notably, inhibitory phosphatases (PTEN, INPP5D) also emerged as candidate substrates. Functional studies revealed proteasome-dependent stabilization of PTEN and INPP5D in GID/CTLH-deficient macrophages with each phosphatase individually exerting an influence on Mtb intracellular survival. Together, our study defines a GID/CTLH-dependent ubiquitylome in macrophages and identifies the complex as a central regulator of metabolism and antimicrobial immunity.

Molecular pathways for learning in the single cell Stentor coeruleus

In arid sandy plantations, stand density critically regulates belowground ecosystems, yet its effects on microbial network complexity, stability, and function are not fully understood. This study examined Pinus sylvestris var. mongholica plantations along a density gradient (Very high density (VHD): 2,450 trees ha-1, High density (HD): 1,633 trees ha-1, Moderate density (MD): 1,067 trees ha-1, Low density (LD): 583 trees ha-1) at two soil depths (0-20, 20-40 cm) in the Otingdag Sandy Land, integrating soil physicochemical, enzymatic, and microbial network analyses. Key findings were: (1) Soil organic carbon, total nitrogen, key enzyme activities [urease (URE), acid phosphatase (PHO), and nitrate reductase (NR)], and ecosystem multifunctionality showed a unimodal response, peaking at medium densities. (2) Microbial responses diverged: bacterial α-diversity changed but composition remained stable, whereas fungal composition was highly density-sensitive. Mid- to low densities promoted more complex, modular, and stable microbial networks. Mantel tests identified pH, URE, and Ammonium nitrogen (NH₄+-N, AN) as key drivers for bacterial phyla, and pH, Soil organic carbon (SOC), Total nitrogen (TN), PHO, and Polyphenol oxidase (PPO) for differentiating Ascomycota and Basidiomycota. (3) Random Forest regression identified microbial network stability as the top predictor of multifunctionality, surpassing diversity. Partial Least Squares Path Modeling (PLS-PM) analysis revealed that stand density enhances multifunctionality primarily by improving the soil environment, with microbial networks acting as environment-dependent regulators. This study demonstrates that moderate stand densities optimize microbial network resilience and ecosystem multifunctionality in sandy plantations, providing a novel perspective from microbial network stability.

Protein tyrosine phosphatase B (PtpB) is a virulence-associated phosphotyrosine phosphatase secreted by Mycobacterium tuberculosis (Mtb), known to disrupt host immune signaling by dephosphorylating key proteins. Targeting PtpB represents a rational strategy for anti-TB drug discovery. This study presents an integrative computational framework for identifying and evaluating small-molecule inhibitors of Mtb PtpB. QSAR models were constructed using four molecular fingerprint types, CDK, PubChem, MACCS, and AtomPairs2DCount, as regression models predicting pIC50 values. Multiple machine learning algorithms were evaluated, with model performance assessed via R2, RMSE, cross-validation, and Y-randomization. SHAP analysis was applied to the top-performing PubChem-SVR model to interpret key structural features. Top-ranked compounds were subjected to molecular docking followed by 250 ns MD simulations to examine binding stability. MM-GBSA and PCA were used for post-simulation analysis. Gene-level interactions were evaluated by comparing predicted compound targets with Mtb-related host genes. Among descriptors, the PubChem-RF model achieved the best performance. SHAP identified PubchemFP417 (alkyne), PubchemFP462 (carboxylic acid), PubchemFP143 (five-membered rings), and PubchemFP34 (sulfur-containing fragments) as major contributors. CHEMBL4635765 showed strong and stable binding within the PTP pocket, while isoxazole carboxylic acid maintained key interactions but with lower stability. Network analysis revealed four shared targets (APP, HDAC8, CACNA1B, pvdQ) and compound-specific links to immune-related genes, including PTPN1 and NFKB1. This integrative computational study combines machine learning, structural modeling, and network pharmacology to provide mechanistic insights into PtpB inhibition and to identify promising chemical scaffolds for future anti-tubercular research. As the analysis is entirely computational, experimental validation will be required to confirm the predicted activities.

Corrigendum to “Aggregation-induced emission Ru(II) complex nanomaterials: Phosphorescent determination of alkaline phosphatase activity and inhibitors” [Microchem. J. 218 (2025) 115191

Microscale engagement of the hemi-immunoreceptor tyrosine-based activation motif-containing receptor Dectin-1 by fungal particles activates Src-family kinases (SFKs) and Syk, drives second-messenger generation, and induces downstream Erk and Akt signaling and proinflammatory responses in macrophages. To avoid inappropriate activation in the absence of a pathogenic threat, macrophages restrict signaling in response to low-valency ligands. To examine how SFKs regulate this sensitivity threshold, we compared signaling induced by pharmacological SFK activation with signaling triggered by depleted zymosan, a high-valency β-glucan particle that engages Dectin-1 to form a phagocytic synapse. We found that particulate engagement of Dectin-1 protected the inhibitory ITIM-associated phosphatase SHIP1 from phosphorylation by SFKs, allowing robust activation of Erk and Akt and proinflammatory induction. In contrast, receptor-independent SFK activation induced phosphorylation of SHIP1 and failed to amplify signaling downstream of PLCγ2 and PI3K. Although multiple SFKs could phosphorylate SHIP1, Lyn uniquely maintained the basal set-point of SHIP1 phosphorylation, thereby keeping PIP3 levels low and suppressing basal Erk and Akt signaling. This Lyn-dependent regulation was essential for suppressing Akt activation and balancing signaling through the Erk and Akt pathways in the absence of a phagocytic synapse. In contrast, antimicrobial responses to particulate stimuli, including second-messenger signaling, Erk/Akt, and proinflammatory outputs, did not strictly require Lyn expression. These findings highlight the unique role of Lyn in limiting spurious proinflammatory signaling and shed light on a mechanism by which macrophages selectively respond to high-valency particulate ligands that override this basal inhibitory program.

Densities of neurofibrillary tangles (NFTs), a major Alzheimer's disease (AD) pathology, display genetic associations with variants in the receptor type protein tyrosine phosphatase D (PTPRD) gene. NFTs are rich in tau protein that is hyperphosphorylated, prominently by the glycogen synthase kinases (GSK) 3α/β. PTPRD dephosphorylates GSK3s, reducing their activities and providing an attractive candidate molecular mechanism for PTPRD/NFT associations. We have used AT-8 and Aβ immunohistochemistry to assess hyperphosphorylated tau/NFT and Aβ/senile plaque pathologies, developed and characterized 3xTg-AD mice with wildtype or reduced PTPRD expression and assessed results of treatments with our (a) PTPRD phosphatase inhibitor pentilludin, (b) lead PTPRD positive allosteric modulator (PAM) quercetin and (c) drug candidate PTPRD PAM active metabolite 6BrQ. Four-month 3xTg-AD/PTPRD +/- mice display AT-8 immunoreactivity in hippocampal neurons, much earlier than 3xTg-AD/PTPRD +/+ mice. There are modest effects of reducing PTPRD expression on densities of Aβ/senile plaque structures assessed at 21 months. 3xTg-AD (but not wildtype C57) mice treated (weeks 6-16) with pentilludin display abundant hyperphosphorylated tau at 4 months. 3xTg-AD/PTPRD+/- mice treated (weeks 6-16) with quercetin or 6BrQ display >50% and >95% reductions in AT-8 immunoreactive hippocampal neuron counts, respectively. These results support roles for PTPRD in AD neurofibrillary pathophysiology and for orally-bioavailable drugs that can be metabolized to 6BrQ to slow development of this pathology.

Among the various virulence factors produced by Mycobacterium tuberculosis, PtpB plays a critical role in regulating the M.tb pathogenesis. Genetic deletion or pharmacological inhibition of PtpB significantly impairs M.tb survival within the host, making it an attractive target for effective antitubercular therapy. Previously, Brunsvicamide B, a cyclic hexapeptide produced by the freshwater cyanobacterium Tychonema sp., was reported to inhibit the enzymatic activity of PtpB. However, the molecular details of the Brunsvicamide B-PtpB interaction remain unresolved, and elucidating this interaction could provide a structural framework for the rational design of peptide-based PtpB inhibitors. In the present study, we characterized the molecular interactions of Brunsvicamide B with PtpB, PtpA, and PstP, identifying key binding parameters responsible for the selective inhibition of PtpB. Pharmacophore models were generated for each interaction, and the complex stability was evaluated using binding energy calculations, RMSD, and RMSF analyses. Comparative analysis with the known PtpB inhibitor OMTS (PDB ID: 2OZ5) revealed that Brunsvicamide B binds to similar functional residues within the catalytic pocket of PtpB. Using the Brunsvicamide B-PtpB complex as a reference model, we screened a library of Anabaenopeptins for their ability to inhibit PtpB and its homologues across the members of the M. tuberculosis complex. This approach led to the identification of three candidate peptides exhibiting favorable safety, bioavailability, and excretion profiles compared to OMTS. Thus, the shortlisted Anabaenopeptins represent promising leads for the development of pan-MTBC PtpB inhibitors with potential application in next-generation TB therapeutics.