• BMB Reports
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• 제34권1호
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• pp.33-38
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• 2001

The human PTK6 (also known as Brk) polypeptide, which is deduced from its full-length cDNA, represents a non-receptor protein tyrosine kinase (PTK). It contains SH3, SH2, and tyrosine kinase catalytic domains that are closely related to Src family members. We generated an antihuman PTK6 antibody by immunizing rabbits with a PTK6-specific oligopeptide conjugated to BSA, which corresponds to 11 amino acid residues near the C-terminus. An immunoblot analysis with the antibody detected an expected 52-kDa band in various mammalian transformed cell lines. Immunoprecipitation and immunoblot analyses demonstrated that PTK6 is phosphorylated on the tyrosine residues) and interacts with approximately a 23-kDa tyrosine-phosphorylated polypeptide (most likely a substrate of PTK6) in breast carcinoma T-47D cells. An immunofluorescence analysis demonstrated that PTK6 is localized throughout the cytoplasm of T-47D cells. These results support a possible role for PTK6 in the intracellular signal transduction through tyrosine phosphorylation.

• Parasites, Hosts and Diseases
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• 제48권4호
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• pp.285-290
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• 2010

Tyrosine kinases are one of the most important regulators for intracellular signal transduction related to inflammatory responses. However, there are no reports describing the effects of tyrosine kinases on neutrophil apoptosis induced by Entamoeba histolytica, In this study, isolated human neutrophils from peripheral blood were incubated with live trophozoites in the presence or absence of tyrosine kinase inhibitors. Entamoeba-induced receptor shedding of CD16 and PS externalization in neutrophils were inhibited by pre-incubation of neutrophils with the broad-spectrum tyrosine kinase inhibitor genistein or the Src family kinase inhibitor PP2. Entamoeba-induced ROS production was also inhibited by genistein or PP2, Moreover, genistein and PP2 blocked the phosphorylation of ERK and p38 MAPK in neutrophils induced by E. histolytica. These results suggest that Src tyrosine kinases may participate in the signaling event for ROS-dependent activation of MAPKs during neutrophil apoptosis induced by E. histolytica.

• The Korean Journal of Physiology and Pharmacology
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• 제16권5호
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• pp.305-312
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• 2012

This study was to determine the effect of exercise on the recovery of dopaminergic neuron loss and muscle atrophy in 6-OHDA-induced hemi Parkinson's disease model. Exercise was loaded twice per day for 30 minutes each time, at 5 days after 6-OHDA lesioning and continued for 16 days using a treadmill. Exercise significantly increased the number of tyrosine hydroxylase positive neuron in the lesioned substantia nigra and the expression level of tyrosine hydroxylase in the striatum compared with the control group. To examine which signaling pathways may be involved in the exercise, the phosphorylation of $GSK3{\beta}$ and ERK were observed in the striatum. In the control group, basal level of $GSK3{\beta}$ phosphorylation was less than in both striatum, but exercise increased it. ERK phosphorylation decreased in the lesioned striatum, but exercise recovered it. These findings suggest that exercise inactivates $GSK3{\beta}$ by phosphorylation which may be involved in the neuroprotective effect of exercise on the 6-OHDA-induced cell death. In the exercise group, weight, and Type I and II fiber cross-sectional area of the contralateral soleus significantly recovered and expression of myosin heavy chain and Akt and ERK phosphorylation significantly increased by exercise. These results suggest that exercise recovers Parkinson's disease induced dopaminergic neuron loss and contralateral soleus muscle atrophy.

• Asian-Australasian Journal of Animal Sciences
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• 제22권2호
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• pp.181-186
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• 2009

In our previous report, we demonstrated that the tyrosine phosphorylation of sperm proteins (TPSP) of guinea pig was associated with capacitation and hyperactivation (CAHA), and $Ca^{2+}$ and ${HCO_3}^-$ were required for the initiation of CAHA and increasing the TPSP. The aim of this study was to further investigate the mechanism underlying the above events. The results showed that addition of cAMP agonists, dibutyryl-cAMP (db-cAMP) and isobutyl-methylxantine (IBMX), to ${HCO_3}^-$ -free medium significantly increased CAHA to the normal level (when sperm were incubated in TALP). Although addition of the cAMP agonists to $Ca^{2+}$-free medium increased CAHA, the percentages of hyperactivated and capacitated sperm were still significantly lower than the normal level. Compared with ${HCO_3}^-$ -free or $Ca^{2+}$-free medium, TPSP was increased when db-cAMP and IBMX were added in the media. H-89, a specific inhibitor of protein kinase A (PKA), inhibited CAHA in a dose-dependent manner and totally blocked TPSP. These results confirm a previous observation that $Ca^{2+}$ and ${HCO_3}^-$ regulated CAHA and TPSP in a cAMP/PKA pathway, and support an interation between TPSP and CAHA of sperm. Besides the cAMP/PKA pathway, $Ca^{2+}$ might have also played a role in regulating CAHA by other pathways since the normal level of CAHA did not recover by adding cAMP agonists in the media.

• IMMUNE NETWORK
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• 제12권4호
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• pp.155-164
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• 2012

It is well established that blocking the interaction of EGFR with growth factors leads to the arrest of tumor growth, resulting in tumor cell death. ER414 is a human monoclonal antibody (mAb) derived by guided selection of the mouse mAb A13. The ER414 exhibited a ~17-fold lower affinity and, as a result, lower efficacy of inhibition of the EGF-mediated tyrosine phosphorylation of EGFR when compared with mAb A13 and cetuximab. We performed a stepwise in vitro affinity maturation to improve the affinity of ER414. We obtained a 3D model of ER414 to identify the amino acids in the CDRs that needed to be mutated. Clones were selected from the phage library with randomized amino acids in the CDRs and substitution of amino acids in the HCDR3 and LCDR1 of ER414 led to improved affinity. A clone, H3-14, with a ~20-fold increased affinity, was selected from the HCDR3 randomized library. Then three clones, ER2, ER78 and ER79, were selected from the LCDR1 randomized library based on the H3-14 but did not show further increased affinities compared to that of H3-14. Of the three, ER2 was chosen for further characterization due to its better expression than others. We successfully performed affinity maturation of ER414 and obtained antibodies with a similar affinity as cetuximab. And antibody from an affinity maturation inhibits the EGF-mediated tyrosine phosphorylation of EGFR in a manner similar to cetuximab.

• Journal of Microbiology and Biotechnology
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• 제29권4호
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• pp.645-650
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• 2019

Brown adipocytes have an important role in the regulation of energy balance through uncoupling protein-1 (UCP-1)-mediated nonshivering thermogenesis. Although brown adipocytes have been highlighted as a new therapeutic target for the treatment of metabolic diseases, such as obesity and type II diabetes in adult humans, the molecular mechanism underlying brown adipogenesis is not fully understood. We recently found that protein tyrosine phosphatase receptor type B (PTPRB) expression dramatically decreased during brown adipogenic differentiation. In this study, we investigated the functional roles of PTPRB and its regulatory mechanism during brown adipocyte differentiation. Ectopic expression of PTPRB led to a reduced brown adipocyte differentiation by suppressing the tyrosine phosphorylation of VEGFR2, whereas a catalytic inactive PTPRB mutant showed no effects on differentiation and phosphorylation. Consistently, the expression of brown adipocyte-related genes, such as UCP-1, $PGC-1{\alpha}$, PRDM16, $PPAR-{\gamma}$, and CIDEA, were significantly inhibited by PTPRB overexpression. Overall, these results suggest that PTPRB functions as a negative regulator of brown adipocyte differentiation through its phosphatase activity-dependent mechanism and may be used as a target protein for the regulation of obesity and type II diabetes.

• Journal of Applied Biological Chemistry
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• 제66권
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• pp.311-319
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• 2023

Caveolin-1 (Cav-1) is the main structural component of the caveolae on the plasma membrane, which regulates various cellular processes, including cell growth, differentiation, and endocytosis. Although a recent study demonstrated that Cav-1 might be involved in diabetes-associated inflammation, its exact role in the intestine was unclear. In this study, we examined the intestinal expression of Cav-1 in diabetic conditions. We also investigated its effect on lipopolysaccharide (LPS)-induced inflammation by expressing this protein in human intestinal Caco-2 cells lacking Cav-1. We observed that increased Cav-1 levels and decreased expression of tight junction proteins affected intestinal permeability in high-fat diet-induced diabetic mice. When Caco-2 cells were treated with LPS, Cav-1 enhanced the NF-κB signaling. Moreover, LPS reduced the expression of tight junction proteins while it increased cell-cell permeability and reactive oxygen species generation in Caco-2 cells and this effect was amplified by cav-1 overexpression. LPS treatment promoted phosphorylation of tyrosine-14 (Y14) on Cav-1, and the LPS-induced NF-κB signaling was suppressed in cells expressing non-phosphorylatable Cav-1 (tyrosine-14 to phenylalanine mutant), which reduced intestinal barrier permeability. These results suggest that Cav-1 expression promotes LPS-induced inflammation in Caco-2 cells, and phosphorylation of Y14 on Cav-1 might contribute to the anti-inflammatory response in LPS-induced NF-κB signaling and cell permeability.

• BMB Reports
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• 제45권12호
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• pp.693-699
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• 2012

Together with protein tyrosine kinases (PTKs), protein tyrosine phosphatases (PTPs) serve as hallmarks in cellular signal transduction by controlling the reversible phosphorylation of their substrates. The human genome is estimated to encode more than 100 PTPs, which can be divided into eleven sub-groups according to their structural and functional characteristics. All the crystal structures of catalytic domains of sub-groups have been elucidated, enabling us to understand their precise catalytic mechanism and to compare their structures across all sub-groups. In this review, I describe the structure and mechanism of catalytic domains of PTPs in the structural context.

• BMB Reports
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• 제50권6호
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• pp.329-334
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• 2017

Protein tyrosine phosphatases (PTPs) play crucial roles in signal transduction and their functional alteration has been detected in many diseases. PTP inhibitors have been developed as therapeutic drugs for diseases that are related to the activity of PTPs. In this study, PTP inhibitor XIX, an inhibitor of CD45 and PTEN, was investigated whether it inhibits other PTPs. Protein tyrosine phosphatase non-receptor type 2 (PTPN2) was selectively inhibited by the inhibitor in a competitive manner. Drug affinity responsive target stability (DARTS) analysis showed that the inhibitor induces conformational changes in PTPN2. Phosphorylation levels of signal transducer and activator of transcription 3 (STAT3) at Tyr-705, a crucial site for STAT3 activation and target site of PTPN2, decreased upon exposure to the inhibitor. Our results suggest that PTP inhibitor XIX might be considered as an effective regulator of PTPN2 for treating diseases related to PTPN2.

• 생명과학회지
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• 제22권3호
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• pp.428-436
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• 2012

단백질 인산화는 세포의 활동을 조절하는 보편적인 과정이다. 브라시노스테로이드(brassinostreoid)에 의해 매개되는 신호전달은 브라시노스테로이드에 의해 활성화된 세포막상의 protein kinase 로부터 인산화되어 있는 전사인자들을 탈인산화하는 연속적인 인산화/탈인산화 과정이다. 브라시노스테로이드에 의해 매개되는 신호전달의 연구는 인산화에 관여하는 kinase 기질상의 아미노산을 밝히고, 그와 관련된 돌연변이체의 표현형을 알아봄으로써 급속하게 발전하였다. BRI1과 BAK1의 자기인산화(autophosphorylation), 상호인산화(transphosphorylation), 타이로신 인산화(tyrosine phosphorylation)를 밝힘으로써 그들의 조절작용을 식물의 생리학적, 발생학적 과정을 더 이해할 수 있는 장이 열렸다. 브라시노스테로이드에 의한 인산화는 수용체에 의해 매개되는 세포 내 함입(endocytosis)과 그에 뒤따르는 수용체의 파괴현상에서도 볼 수 있다. 인산화/탈인산화 과정에 관련하여 브라시노스테로이드에 의해 매개되는 신호전달은 더 연구할 여지가 많이 남아 있다. 이 총설은 단백질의 인산화/탈인산화 과정을 통한 브라시노스테로이드의 신호전달 연구의 최근 상황을 기술하였다.