Phospho-PTPN1 (Ser50) Antibody
Description
Antigen Overview: PTP1B and Ser50 Phosphorylation
PTP1B, encoded by the PTPN1 gene, is a non-receptor protein tyrosine phosphatase involved in insulin/leptin signaling, growth factor regulation, and integrin signaling . Its structure includes:
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A catalytic phosphatase domain.
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C-terminal hydrophobic residues for endoplasmic reticulum (ER) membrane anchoring.
Phosphorylation at Ser50 modulates PTP1B activity:
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Functional Impact: Conflicting reports suggest phosphorylation at Ser50 either:
Role in Metabolic Regulation
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PTP1B dephosphorylates IR and IRS proteins, acting as a negative regulator of insulin signaling .
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Ser50 phosphorylation by Akt creates a feedback loop to attenuate PTP1B activity, enhancing insulin sensitivity .
Disease Implications
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Obesity/Diabetes: Ptp1b⁻/⁻ mice exhibit leanness and improved glucose homeostasis .
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Cancer: PTP1B promotes oncogenic signaling (e.g., Src activation) in breast cancer models .
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Inflammation: PTP1B inhibitors reduce acute lung injury (ALI) and sepsis by suppressing neutrophil migration and NETosis .
Post-Translational Modifications (PTMs) at Ser50
Applications in Research
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Mechanistic Studies: Elucidating PTP1B’s dual role in metabolism and cancer .
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Therapeutic Development: Validating PTP1B inhibitors (e.g., MSI-1436, DPM-1003) in preclinical models of ALI and sepsis .
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Disease Biomarkers: Detecting phosphorylation status in insulin resistance or tumor samples .
Key Research Findings
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Neutrophil Regulation: PTP1B inhibitors suppress CXCR4/PI3Kγ/AKT/mTOR signaling in neutrophils, reducing lung injury in TRALI models .
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Autoimmunity: PTP1B deficiency in B cells elevates CD40/BAFF-R signaling, increasing autoantibody production .
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Structural Insights: Oxidation at Cys215 induces conformational changes, reversibly inactivating PTP1B .
Product Specs
Target Background
- The Cav-2beta isoform, produced by alternative translation initiation, desensitizes the insulin receptor (IR) via dephosphorylation by PTP1B, followed by endocytosis and lysosomal degradation of IR, leading to insulin resistance. PMID: 29604334
- High-resolution PTP1B inhibition profiling combined with HPLC-HRMS-SPE-NMR for identification of PTP1B inhibitors from Miconia albicans. PMID: 30018269
- The expression of PTP1B in skeletal muscle was found to be elevated after caloric restriction and prolonged exercise. PMID: 28729389
- Preliminary data indicate that heterozygous or homozygous deletion of PTPN1 increases the severity of MPN in Jak2V617F-knock-in mice. Further research will determine whether the loss of PTPN1 collaborates with the JAK2V617F mutation in the pathogenesis of MPN. PMID: 28111468
- Our research demonstrated a novel oncogenic mechanism of PTP1B involving its impact on PITX1/p120RasGAP in colorectal carcinoma (CRC). Regorafenib inhibited CRC survival by preserving PTP1B-dependent PITX1/p120RasGAP downregulation. PTP1B may serve as a potential biomarker for predicting regorafenib effectiveness and a potential therapeutic target for CRC. PMID: 27752061
- This study highlights a crucial role for Nck1 in fine-tuning IRE1alpha expression and signaling that regulate PTP1B expression and subsequent activation of the PI3K-Akt pathway in HepG2 cells. PMID: 28455143
- In conclusion, HDAC6 might enhance aggressive melanoma cell progression by interacting with PTPN1, independent of its histone modifying activity. PMID: 29278704
- PTP1B is widely expressed in the human breast gland, with the highest expression observed in myoepithelial cells and fibroblasts. Inhibition of PTP1B in D492 and HMLE affects cell-cell adhesion and induces anoikis-like effects. PMID: 28492548
- The results of the current study emphasize the expression quantitative trait loci enrichment and pleiotropy in psoriasis and schizophrenia, and also suggest a potential key role of the PTPN1 gene in the etiology of psoriasis. PMID: 27976820
- The identified structural fragments, crucial for PTP1B inhibition, were determined using a naive Bayesian method and can be further exploited to design new molecules around the identified scaffolds. The descriptive and predictive modeling strategy employed in this study is capable of identifying PTP1B inhibitors from extensive compound libraries. PMID: 28006676
- The obtained results not only reveal the unbinding mechanism of IRK-PTP1B complexes, derived from pulling force profile, number of hydrogen bonds, and interaction energy between IRK and PTP1Bs, but also describe how PTP1B's point mutations can variably alter its binding affinity towards IRK. PMID: 28707052
- These findings suggest that inhibiting PTP1B activity is a promising novel therapeutic target in the treatment of colorectal cancer and the prevention of metastasis. PMID: 26942883
- The molecular mechanisms by which PTP1B and TC-PTP loss cooperate with other genetic aberrations need to be elucidated to develop more effective therapeutic strategies. PMID: 26817397
- The findings demonstrate that reduced PTP1B responses contribute to disease symptoms, partly by enhancing S100A9 expression during viral-associated chronic obstructive pulmonary disease exacerbations. PMID: 26813343
- Both the rs2904268 C>G CG and GG genotype frequencies were significantly higher in the ESCC group compared to the control group (both p < 0.05). However, the genotype frequencies of rs2230605A>G and rs16995309 C>T were similar between the ESCC and control groups. These results suggest that the PTPN1 gene polymorphism rs2904268 is associated with susceptibility to Esophageal Squamous Cell Carcinoma in Inner Mongolia. PMID: 28287835
- Data suggests that TrxR1 and NADPH directly protect PTP1B from inactivation by oxidation; this protection is independent of TRX1 and PRX2; this protection is blocked by auranofin (an inhibitor of TrxR1) and requires an intact selenocysteine residue in TrxR1. (TrxR1 = thioredoxin reductase 1; PTP1B = protein tyrosine phosphatase, non-receptor type 1; TRX1 = thioredoxin-1; PRX2 = paired related homeobox 2 protein) PMID: 28684416
- Regulation of platelet-activating factor-mediated PTP1B activation by a Janus kinase 2/calpain pathway has been reported. PMID: 28686728
- PTP1B was overexpressed in over 70% of breast cancer tissues, correlating with patients with estrogen receptor (ER)-negative, progesterone receptor (PR)-negative, and human epidermal growth factor receptor 2 (HER2)-positive tumors. The data also indicated that both tumor size and lymph node metastasis were significantly higher in patients with a higher level of PTP1B. PMID: 28378571
- PTP1B utilizes conformational and dynamic allostery to regulate its activity. Both conformational rigidity and dynamics are essential for controlling protein activity. PMID: 28212750
- PTP1BDelta6 is a positive regulator of JAK/STAT signaling in classical Hodgkin lymphoma cells. (PMID: 28082443
- In PTP1B and VHR, two new allosteric clusters were identified in each enzyme. PMID: 28625849
- The PTP1BDeltaE6 mutant expressed in colon and thyroid tumors and another PTP1BDeltaE5 mutant expressed in colon tumor were studied in more detail. PMID: 27855221
- The PTP1B/RNF213/alpha-KGDD pathway is crucial for the survival of HER2(+) breast cancer, and possibly other malignancies, in the hypoxic tumor microenvironment. PMID: 27323329
- Data show for the first time a calpain/PTP1B/VEGFR2 negative feedback loop in the regulation of VEGF-induced angiogenesis. Modulation of local PTP1B and/or calpain activities may prove beneficial in the treatment of impaired wound healing in diabetes. PMID: 27872190
- Data show that cichoric acid (CHA) is a competitive inhibitor that binds in the active site of protein tyrosine phosphatase 1B (PTP1B), and chlorogenic acid (CGA), while a noncompetitive inhibitor, binds in the second aryl phosphate binding site, rather than the predicted benzfuran binding pocket. PMID: 27959494
- Stimulation of the DOCK180 pathway represents an alternative mechanism of PTP1B inhibitor-stimulated endothelial cell motility, which does not require concomitant VEGFR2 activation as a prerequisite. PMID: 27052191
- Overexpression of PTP1B is associated with breast neoplasms. PMID: 27465552
- The analysis revealed three candidate genes, GSK3B, PTPN1, and STX4, that are differentially expressed in study subjects. GSK3B was highly significant in Ps-T2D (P=0.00018, FR=-26.6), followed by Ps (P=0.0028, FR=-14.5) and T2D groups (P=0.032, FR=-5.9). PTPN1 showed significant association only with PS-T2D (P=0.00027, FR=-8.5). STX4 showed significant association with both Ps (P=0.0002, FR=-20) and Ps-T2D (P=0.0016, FR=-11.2). PMID: 27530212
- PTP1B directly regulates STAT5 phosphorylation, and its activation via the cAMP/PKA pathway downstream of the 5-HT7 receptor is involved in the suppression of beta-casein expression in MCF-12A cells. PMID: 27016479
- Upon EGF stimulation, complexes containing Mena and PTP1B are recruited to the EGFR, causing receptor dephosphorylation and leading to decreased motility responses. PMID: 26337385
- PTP1B dephosphorylates PITX1 to weaken its protein stability and the transcriptional activity for p120RasGAP gene expression. PMID: 26840794
- Our data indicate that by targeting PTP1B, miR-744 plays a feed-forward role in regulating the type I IFN signaling pathway. PMID: 26259828
- O-GlcNAcylation of PTP1B can influence insulin signal transduction by modulating its own phosphatase activity, which participates in the process of hepatic insulin resistance. PMID: 26402673
- Blockade of the interaction between Bcr-Abl and PTB1B by small molecule SBF-1 overcomes imatinib-resistance of K562 cells. PMID: 26721204
- The multiple roles of PTP1B in liver diseases will be presented, with respect to liver regeneration, drug-induced liver disease, non-alcoholic fatty liver disease (NAFLD), and hepatocellular carcinoma. [REVIEW] PMID: 26299811
- PTP1B amplification predicts poor survival of gastric cancer patients. PMID: 25590580
- The hydrogen bond and energy decomposition analyses indicate that, apart from residue 181, point mutations influence the interactions of the substrate with several residues in the active site of PTP1B. PMID: 24865376
- Nck1 depletion induces activation of the PI3K/Akt pathway by attenuating PTP1B protein expression. PMID: 25398386
- Findings suggest that the second sphere residues of the PTP1B active site may play significant roles in PTP-mediated function in both normal and diseased states. PMID: 25450460
- PTP1B sub-cellular location and tight control of enzymatic activity by diverse post-translational modifications illustrate the dynamic regulation of this enzyme and its ability to modulate numerous signaling pathways, likely in a cell/tissue- and stimulus-dependent manner, with high specificity and precision. [Review] PMID: 25263014
- FGFR3 regulation by PTPN1 and PTPN2 depends on FGFR3 localization and A-loop sequence. PMID: 25311528
- Data show a relationship between the dephosphorylation rate of different factors in signaling pathways related to cell proliferation and PTP1B activation. PMID: 25668475
- Fumosorinone increased glucose uptake and improved insulin resistance by down-regulating the expression of PTP1B and activating the insulin signaling pathway, suggesting that it may possess antidiabetic properties. PMID: 25796170
- M3R preserves the endothelial barrier function through a mechanism potentially maintaining PTP1B activity, keeping the adherens junction proteins (AJPs) dephosphorylated. PMID: 24393526
- Data indicate that protein tyrosine phosphatase 1B (PTP1B) association with calnexin is ubiquitin conjugating enzyme 9 (UBC9)-dependent. PMID: 25586181
- PTP1B can promote non-small cell lung cancer proliferation and metastasis by activating src and subsequently ERK1/2. PMID: 25617799
- Protein tyrosine phosphatase 1B is regulated with a conformational rearrangement in helix alpha7, disrupting the triangular interaction among helix alpha7, helix alpha3, and loop11. PMID: 24831294
- PTP1B is a novel physiological regulator of TrkB, and enhanced BDNF/TrkB signaling may contribute to the beneficial metabolic effects of PTP1B deficiency. PMID: 25288805
- Nox4-mediated redox regulation of PTP1B serves as a modulator, in part through coronin-1C, of the growth and migration of glioblastoma cells. PMID: 24239742
- PTP1B plays a role in the modulation of insulin signaling by acetaminophen in hepatocytes. PMID: 25204659
Q&A
Fundamental Aspects of PTP1B and Ser50 Phosphorylation
Q: What is the biological significance of PTP1B phosphorylation at Ser50?
Phosphorylation of PTP1B at Ser50 represents a critical regulatory mechanism that modulates its phosphatase activity. When Akt phosphorylates PTP1B at Ser50, it impairs PTP1B's ability to dephosphorylate the insulin receptor, creating a positive feedback mechanism for insulin signaling . This phosphorylation event serves as a molecular switch that reduces PTP1B activity toward its substrates, particularly the insulin receptor and IRS-1, thereby enhancing downstream signaling pathways . The Ser50 site is located within an Akt consensus motif (RYRDVS50), making it a direct target for Akt-mediated regulation in response to insulin stimulation .
Q: How does phosphorylation at Ser50 mechanistically affect PTP1B function?
Phosphorylation at Ser50 appears to impair PTP1B function through multiple mechanisms:
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Reduced substrate binding: Experiments with phosphomimetic mutants (PTP1B-S50D) demonstrate decreased binding to substrates like the insulin receptor
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Decreased catalytic activity: In vitro phosphatase assays show that Ser50 phosphorylation directly reduces enzymatic activity toward phosphorylated substrates
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Substrate-specific effects: The inhibitory impact may be more pronounced for insulin receptor dephosphorylation than for other substrates like IRS-1
This mechanism represents a regulatory circuit that potentiates insulin signaling by inhibiting a negative regulator (PTP1B).
Experimental Applications and Technical Considerations
Q: What experimental applications are suitable for Phospho-PTP1B (Ser50) antibodies?
Phospho-PTP1B (Ser50) antibodies can be utilized in multiple experimental applications:
The antibody is particularly valuable for studying insulin signaling pathways, Akt-mediated phosphorylation events, and the regulatory mechanisms of PTP1B activity in cellular contexts .
Q: How can I verify the specificity of Phospho-PTP1B (Ser50) antibody in my experiments?
To verify antibody specificity, implement these methodological approaches:
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Phosphatase treatment control: Treat one sample with lambda phosphatase before immunoblotting. The signal should significantly decrease in treated samples
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Mutant controls: Use cells expressing PTP1B-S50A (cannot be phosphorylated) as negative controls and PTP1B-S50D (phosphomimetic) as positive controls
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Calyculin A treatment: As a positive control, treat cells with calyculin A (a phosphatase inhibitor) to enhance phosphorylation signals, which should appear as a 50 kDa doublet in Western blots
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Akt inhibition: Use specific Akt inhibitors or PI3K inhibitors like wortmannin to reduce Ser50 phosphorylation, which should decrease the signal
Advanced Research Methodologies
Q: What controls are essential when using Phospho-PTP1B (Ser50) antibody to study insulin signaling?
When studying insulin signaling with Phospho-PTP1B (Ser50) antibody, include these critical controls:
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Basal vs. insulin-stimulated conditions: Compare samples from serum-starved cells with and without insulin stimulation (typically 100 nM insulin for 10 minutes)
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PI3K/Akt pathway inhibition: Include samples pre-treated with wortmannin (100 nM) to block the pathway upstream of Akt
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Total PTP1B control: Always blot for total PTP1B (using a non-phospho-specific antibody) to normalize phospho-signals
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Time course experiment: Include multiple time points after insulin stimulation to capture phosphorylation dynamics
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Orthophosphate labeling: For metabolic labeling experiments, use [32P]-orthophosphate (75 μCi/ml in KRB buffer) for 4 hours before insulin stimulation
Q: What methodologies can detect changes in PTP1B activity following Ser50 phosphorylation?
Several methodological approaches can assess PTP1B activity changes:
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In vitro phosphatase assays:
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Phosphotyrosine profiling:
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PTP1B-substrate interaction studies:
Cell Treatments and Detection Optimization
Q: What cell treatments can enhance detection of phospho-Ser50 PTP1B?
To optimize detection of phospho-Ser50 PTP1B in experimental systems:
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Insulin stimulation: Treat serum-starved cells with 100 nM insulin for 10 minutes to activate the PI3K/Akt pathway
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Calyculin A treatment: This phosphatase inhibitor enhances detection by preventing dephosphorylation of Ser50, resulting in a characteristic doublet pattern in Western blots
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Constitutively active Akt: Transfection with myristoylated Akt (Akt-myr) can drive Ser50 phosphorylation independently of upstream signals
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Combined approach: For maximum detection, combine insulin stimulation with phosphatase inhibition
Q: How does phosphorylation at Ser50 compare with other known PTP1B post-translational modifications?
PTP1B undergoes multiple modifications that distinctly regulate its function:
Ser50 is considered the major serine phosphorylation site compared to Ser242 and Ser243 . Unlike tyrosine phosphorylation sites that may enhance certain protein-protein interactions, Ser50 phosphorylation appears primarily inhibitory toward phosphatase activity.
Research Applications in Disease Models
Q: How can Phospho-PTP1B (Ser50) antibodies contribute to studies of metabolic disorders?
Phospho-PTP1B (Ser50) antibodies offer valuable research tools for metabolic disorder studies:
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Insulin resistance mechanisms:
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Therapeutic target validation:
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Cell-based screening approaches:
Q: What is the relationship between PTP1B Ser50 phosphorylation and cancer research?
The study of PTP1B Ser50 phosphorylation in cancer presents several research opportunities:
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PTP1B may play a tumor-promoting role in certain cancers (e.g., prostate cancer), suggesting PTP1B inhibition could be therapeutically valuable
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Since Akt is frequently hyperactivated in many cancers, the Akt-mediated phosphorylation of PTP1B at Ser50 may represent a mechanism through which cancer cells modulate signaling pathways
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Methodological approaches to study this relationship should include:
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Phospho-PTP1B (Ser50) antibodies can be used to evaluate whether PTP1B inhibitors affect this regulatory phosphorylation in cancer models
