Protein tyrosine phosphatases (PTPs, EC 3.1.3.48) are key regulators of cellular processes, with the catalytic activity attributed to the conserved motif (H/V)CX₅R(S/T), where cysteine and arginine residues are critical. Previous studies revealed that alternative splicing of extracellular phosphatase mRNA precursors in Metarhizium anisopliae generated two distinct transcripts, with the longer sequence containing a novel HCPTPMLS motif resembling PTP signatures but lacking the arginine residue. To identify the novel signature motif and characterize its enzymatic properties, we heterologously expressed and purified both proteins in Pichia pastoris and comprehensively characterized their enzymatic properties. The protein containing the HCPTPMLS motif (designated as L-protein) exhibited the highest activity at pH 5.5 and a strong preference for pTyr substrates. Its phosphatase activity was inhibited by Ag⁺, Zn²⁺, Cu²⁺, molybdate, and tungstate, but enhanced by Ca²⁺ and EDTA. AcP101 (lacking HCPTPMLS) showed the maximal activity at pH 6.5 and a strong preference toward pNPP (P < 0.05), with the activity inhibited by NaF and tartrate, but enhanced by Mg²⁺ and Mn²⁺. Functional analysis confirmed that the L-protein retained the PTP activity despite the absence of arginine in its signature motif, while AcP101 functioned as an acid phosphatase. This study provides the first functional validation of an arginine-deficient PTP motif, expanding the definition of PTP signature motifs and offering new insights for phosphatase classification.
Metarhizium/genetics* ; Protein Tyrosine Phosphatases/chemistry* ; Amino Acid Motifs ; Recombinant Proteins/biosynthesis* ; Amino Acid Sequence ; Pichia/metabolism* ; Fungal Proteins/chemistry* ; Substrate Specificity ; Saccharomycetales

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 IC being 5.11 ± 0.53 μmol·L, and acted as a noncompetitive inhibitor based on kinetic analysis.
4-Butyrolactone ; analogs & derivatives ; chemistry ; isolation & purification ; pharmacology ; Animals ; Aspergillus ; chemistry ; Chemistry Techniques, Analytical ; Dipeptides ; chemistry ; isolation & purification ; pharmacology ; Enzyme Inhibitors ; chemistry ; isolation & purification ; pharmacology ; Indoles ; chemistry ; isolation & purification ; pharmacology ; Molecular Structure ; Mycobacterium tuberculosis ; drug effects ; Peptides ; chemistry ; isolation & purification ; pharmacology ; Polyketides ; chemistry ; isolation & purification ; pharmacology ; Porifera ; microbiology ; Protein Tyrosine Phosphatases ; chemistry

Protein tyrosine phosphatase 1B (PTP1B) has led to an intense interest in developing its inhibitors as anti-diabetes, anti-obesity and anti-cancer agents. The fruits of Rubus chingii (Chinese raspberry) were used as a kind of dietary traditional Chinese medicine. The methanolic extract of R. chingii fruits exhibited significant PTP1B inhibitory activity. Further bioactivity-guided fractionation resulted in the isolation of three PTP1B inhibitory ursane-type triterpenes: ursolic acid (1), 2-oxopomolic acid (2), and 2α, 19α-dihydroxy-3-oxo-urs-12-en-28-oic acid (3). Kinetics analyses revealed that 1 was a non-competitive PTP1B inhibitor, and 2 and 3 were mixed type PTP1B inhibitors. Compounds 1-3 and structurally related triterpenes (4-8) were further analyzed the structure-activity relationship, and were evaluated the inhibitory selectivity against four homologous protein tyrosine phosphatases (TCPTP, VHR, SHP-1 and SHP-2). Molecular docking simulations were also carried out, and the result indicated that 1, 3-acetoxy-urs-12-ene-28-oic acid (5), and pomolic acid-3β-acetate (6) bound at the allosteric site including α3, α6, and α7 helix of PTP1B.
Enzyme Inhibitors ; chemistry ; metabolism ; Fruit ; chemistry ; Humans ; Kinetics ; Methanol ; chemistry ; Molecular Docking Simulation ; Molecular Structure ; Plant Extracts ; chemistry ; Protein Binding ; Protein Tyrosine Phosphatase, Non-Receptor Type 1 ; antagonists & inhibitors ; metabolism ; Protein Tyrosine Phosphatases ; antagonists & inhibitors ; Rubus ; chemistry ; Structure-Activity Relationship ; Triterpenes ; chemistry ; metabolism

Protein phosphorylation on tyrosine has emerged as a key device in the control of numerous cellular functions in bacteria. In this article, we review the structure and function of bacterial tyrosine kinases and phosphatases. Phosphorylation is catalyzed by autophosphorylating adenosine triphosphate-dependent enzymes (bacterial tyrosine (BY) kinases) that are characterized by the presence of Walker motifs. The reverse reaction is catalyzed by three classes of enzymes: the eukaryotic-like phosphatases (PTPs) and dual-specific phosphatases; the low molecular weight protein-tyrosine phosphatases (LMW-PTPs); and the polymerase-histidinol phosphatases (PHP). Many BY kinases and tyrosine phosphatases can utilize host cell proteins as substrates, thereby contributing to bacterial pathogenicity. Bacterial tyrosine phosphorylation/dephosphorylation is also involved in biofilm formation and community development. The Porphyromonas gingivalis tyrosine phosphatase Ltp1 is involved in a restraint pathway that regulates heterotypic community development with Streptococcus gordonii. Ltp1 is upregulated by contact with S. gordonii and Ltp1 activity controls adhesin expression and levels of the interspecies signal AI-2.
Bacteria ; enzymology ; Bacterial Proteins ; genetics ; metabolism ; Biofilms ; growth & development ; Gene Expression Regulation, Bacterial ; Host-Pathogen Interactions ; Phosphorylation ; Polysaccharides, Bacterial ; biosynthesis ; Porphyromonas gingivalis ; enzymology ; Protein Processing, Post-Translational ; Protein Structure, Tertiary ; Protein Tyrosine Phosphatases ; chemistry ; genetics ; metabolism ; Protein-Tyrosine Kinases ; chemistry ; genetics ; metabolism ; Quorum Sensing ; Signal Transduction ; Streptococcus gordonii ; enzymology ; Virulence Factors ; metabolism

BACKGROUND AND OBJECTIVEPhosphatase of regenerating liver-3 (PRL-3) is a newly identified protein-tyrosine phosphatase, which belongs to phosphatase of regenerating liver family, and plays a role in promoting tumor metastasis; Ras homologue C (RhoC) belongs to Rho subfamily of small-molecule G protein superfamily. However, the mechanisms of PRL-3 and RhoC are unknown. The aim of this study is to investigate the expressions of PRL-3 and RhoC proteins and their correlation to invasion and metastasis of non-small cell lung cancer (NSCLC), which may provide experiment evidence of the mechanism of PRL-3 in tumorigenesis and tumor-development.

METHODSImmunohistochemical staining was used to detect the expressions of PRL-3 and RhoC in NSCLC in 92 cases, and statistical methods were used to analyse statistical significances of their expressions in different groups and their correlation.

RESULTSThe positive rates of PRL-3 and RhoC expressions in NSCLC were 69.6% (64/92) and 73.9% (68/92), respectively, and the expressions of PRL-3 and RhoC were closely correlated with TNM stage and lymphatic metastasis and pleural metastasis (P < 0.01), and they were correlated with each other (r = 0.754, P < 0.001).

CONCLUSIONThe expressions of PRL-3 and RhoC are higher in the higher TNM stage and lymphatic metastasis and pleural metastasis cases, and closely correlate with each other in NSCLC, which suggests that PRL-3 and RhoC might be in the same signal pathway and PRL-3 might promote the distant metastasis of cancer cell by RhoC and downstream factors.

Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; chemistry ; pathology ; Female ; Humans ; Immunohistochemistry ; Lung Neoplasms ; chemistry ; pathology ; Male ; Middle Aged ; Neoplasm Metastasis ; Neoplasm Proteins ; analysis ; genetics ; physiology ; Neoplasm Staging ; Protein Tyrosine Phosphatases ; analysis ; genetics ; physiology ; rho GTP-Binding Proteins ; analysis ; genetics ; physiology ; rhoC GTP-Binding Protein

To obtain higher potency and specificity, a series of 7-alkoxy analogues of illudalic acid was synthesized on the base of structure-activity relationship (SAR). All of these compounds exhibited submicromolar inhibition of the enzyme when tested against human leukocyte common antigen-related phosphatase (LAR) (for example, for 15e, IC50 = 180 nmol x L(-1)). They represent the most potent small-molecule inhibitors of LAR so far. These analogues also display excellent selectivity for LAR over other protein tyrosine phosphatases (PTPs) except for the highly homologous PTPsigma. The compound 15f is of 120-fold selectivity for LAR versus PTP-1B inhibition. The development of potent enzyme-specific inhibitors is so important that they may serve both as tools to study the role of LAR and as therapeutic agents for treatment of type II diabetes.
Coumarins ; chemical synthesis ; chemistry ; pharmacology ; Enzyme Inhibitors ; chemical synthesis ; chemistry ; pharmacology ; Inhibitory Concentration 50 ; Molecular Structure ; Receptor-Like Protein Tyrosine Phosphatases, Class 2 ; antagonists & inhibitors ; Structure-Activity Relationship

OBJECTIVETo study the effects of androgen on the expression of phosphacan and NG2 proteoglycan (NG2) and neurite regeneration in neonatal rats with hypoxic-ischemic brain damage (HIBD) and the potential mechanism underlying the protective effect of androgen against HIBD.

METHODSOne hundred and twenty neonatal Sprague-Dawley rats were randomly divided into three groups: sham-operated, HIBD and androgen treatment. HIBD was induced by the ligation of left common carotid artery and hypoxia exposure. The androgen treatment group rats were injected with testosterone propionate (25 mg/kg) immediately after HIBD. Phosphacan and NG2 expression in the cortex and the hippocampus was detected with the immunohistochemical method 24 and 72 hrs and 7 and 10 days after hypoxia-ischemia (HI). The ultrastructure and neurite regeneration of neurons in the cortex and the hippocampus were observed under a transmission electron microscope.

RESULTSThe neurite regeneration was obvious in the sham-operated group, but seldom in the HIBD group. The androgen treatment group showed increased neurite regeneration compared with the HIBD group. There were fewer phosphacan and NG2 positive cells in the cortex and the hippocampus in the sham-operated group. Phosphacan and NG2 expression in the cortex and the hippocampus was observed at 24 hrs, increased at 72 hrs, and peaked at 7 days after HI in the HIBD group and remained at a higher expression 10 days after HI than in the sham-operated group. The levels of phosphacan and NG2 expression in the cortex and the hippocampus in the androgen treatment group were significantly reduced compared with those in the HIBD group 24 and 72 hrs and 7 and 10 days after HI (P<0.01).

CONCLUSIONSPhosphacan and NG2 may be important inhibitory factors for neurite regeneration following HIBD in neonatal rats. The neuroprotection of androgen against neonatal HIBD is produced possibly through an inhibition of phosphacan and NG2 expression.

Animals ; Animals, Newborn ; Antigens ; analysis ; Brain Chemistry ; drug effects ; Female ; Hypoxia-Ischemia, Brain ; physiopathology ; Immunohistochemistry ; Male ; Microscopy, Electron, Transmission ; Nerve Regeneration ; drug effects ; Neurites ; physiology ; ultrastructure ; Proteoglycans ; analysis ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptor-Like Protein Tyrosine Phosphatases, Class 5 ; analysis ; Testosterone Propionate ; pharmacology

OBJECTIVETo obtain the entire coding sequence of human PRL-3 gene and construct its prokaryotic expression vector.

METHODSWith total RNA extracted from the human colorectal carcinoma cell line SW480 as the template, PRL-3 gene was amplified by RT-PCR with primers designed according to the published sequence of GenBank, and the product was inserted into pGEM-T Easy vector. The recombinant plasmid pGEM-T-PRL-3 was identified by restriction endonuclease analysis and DNA sequencing. After digestion with restriction endonuclease, PRL-3 gene was cloned into the multicloning sites of the prokaryotic expression vector pGEX-4T-1.

RESULTS AND CONCLUSIONThe entire coding region of human PRL-3 gene was cloned, and the recombinant pGEX-4T-1-PRL-3 vector was successfully constructed and expressed, which may provide the basis for further study of the relationship between human colorectal carcinoma and PRL-3 gene.

Base Sequence ; Cell Line, Tumor ; Cloning, Molecular ; DNA, Complementary ; chemistry ; genetics ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Genetic Vectors ; genetics ; Humans ; Molecular Sequence Data ; Neoplasm Proteins ; genetics ; metabolism ; Protein Tyrosine Phosphatases ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; metabolism ; Sequence Analysis, DNA

BACKGROUNDChina is one of the countries with the highest incidence of H. pylori and more than 9090 isolates possessed the cagA gene. This study was to evaluate the biological activity of the H. pylori virulence factor cagA isolated from Chinese patients.

METHODScagA DNA fragments were amplified from the genomic DNA and subsequently cloned into the mammalian expression vector for cell transfection and DNA sequencing. cagA protein, phosphorylated-tyrosine cagA and the complex of cagA precipitated with SHP-2 were identified respectively by western blot in the crude cell lysate from conditionally immortalized gastric epithelial cells at 48 hours after transfection with cagA DNA. In addition, the ability of induction of scattering phenotype was examined after transient expression of cagA in AGS cells.

RESULTSThe C-terminal half of cagA contained only one repeated sequence and three tandem five-amino-acid motifs glutamic acid-proline-isoleucine-tyrosine-alanine (EPIYA). Moreover, the amino acid sequence of D2 region in repeated sequence was aspartic acid-phenylanaline-aspartic acid (D-F-D) which was significantly distinguished from the three repeated sequences and aspartic acid-aspartic adid-leucine (D-D-L) in the western standard strain NCTC11637. Western blot revealed that cagA became phosphorylated in tyrosine site and bound with SHP-2 after transient expression of cagA DNA in gastric epithelial cells. Transient expression of cagA in AGS cells showed that cagA was able to induce the elongation phenotype although to a lesser extent than western strains.

CONCLUSIONScagA perturbs cell signaling pathways by binding with SHP-2. However, significant difference exists in amino acid sequence and biological function of cagA in Chinese compared with those of western countries.

Amino Acid Sequence ; Antigens, Bacterial ; chemistry ; physiology ; Bacterial Proteins ; chemistry ; physiology ; Blotting, Western ; Cells, Cultured ; Gastric Mucosa ; Humans ; Intracellular Signaling Peptides and Proteins ; Molecular Sequence Data ; Phenotype ; Phosphorylation ; Protein Tyrosine Phosphatase, Non-Receptor Type 11 ; Protein Tyrosine Phosphatases ; metabolism ; Repetitive Sequences, Amino Acid ; Signal Transduction