• Molecules and Cells
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• 제28권3호
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• pp.195-200
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• 2009

Protein tyrosine kinases (PTKs) play a central role in the modulation of a wide variety of cellular events such as differentiation, proliferation and metabolism, and their unregulated activation can lead to various diseases including cancer and diabetes. PTKs represent a diverse family of proteins including both receptor tyrosine kinases (RTKs) and non-receptor tyrosine kinases (NRTKs). Due to the diversity and important cellular roles of PTKs, accurate classification methods are required to better understand and differentiate different PTKs. In addition, PTKs have become important targets for drugs, providing a further need to develop novel methods to accurately classify this set of important biological molecules. Here, we introduce a novel statistical model for the classification of PTKs that is based on their structural features. The approach allows for both the recognition of PTKs and the classification of RTKs into their subfamilies. This novel approach had an overall accuracy of 98.5% for the identification of PTKs, and 99.3% for the classification of RTKs.

• Journal of Microbiology and Biotechnology
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• 제24권2호
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• pp.152-159
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• 2014

The regulation of protein tyrosine phosphorylation is mediated by protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs) and is essential for cellular homeostasis. Co-expression of PTKs with PTPs in Pichia pastoris was used to facilitate the expression of active PTKs by neutralizing their apparent toxicity to cells. In this study, the gene encoding phosphatase PTP1B with or without a blue fluorescent protein or peroxisomal targeting signal 1 was cloned into the expression vector pAG32 to produce four vectors. These vectors were subsequently transformed into P. pastoris GS115. The tyrosine kinases EGFR-2 and $PDGFR{\beta}$ were expressed from vector pPIC3.5K and were fused with a His-tag and green fluorescent protein at the N-terminus. The two plasmids were transformed into P. pastoris with or without PTP1B, resulting in 10 strains. The EGFR-2 and $PDGFR{\beta}$ fusion proteins were purified by $Ni^{2+}$ affinity chromatography. In the recombinant P. pastoris, the PTKs co-expressed with PTP1B exhibited higher kinase catalytic activity than did those expressing the PTKs alone. The highest activities were achieved by targeting the PTKs and PTP1B into peroxisomes. Therefore, the EGFR-2 and $PDGFR{\beta}$ fusion proteins expressed in P. pastoris may be attractive drug screening targets for anticancer therapeutics.

• 대한의생명과학회지
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• 제13권2호
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• pp.119-125
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• 2007

It is widely known that protein tyrosine kinases (PTKs) are involved in controlling many biological processes such as cell growth, differentiation, proliferation, survival and apoptosis. An $\alpha3\beta4$ subunit combination acts as a major functional acetylcholine receptor (nAChRs) in male rat major pelvic ganglion (MPG) neurons, and their activation induces fast inward currents and intracellular calcium increases. Recently it has been reported that the activity of acetylcholine receptors (AChRs) in some neurons can be negatively regulated by PTKs. However, the exact mechanism of regulation of nAChRs by PTKs is poorly understood. Therefore, we examined the potential role particular in nAChR by PTK using electrophysiology and calcium imaging in male rat MPG neurons. ACh induced inward currents and $(Ca^{2+})_i$ increases in MPG neurons, concomitantly. These responses were inhibited by more than 90% in $Na^+$- or $Ca^{2+}$- free solution. $\alpha$-conotoxin AuIB, a selective $\alpha3\beta4$ nAChR blocket, inhibited ACh-induced inward currents. Genistein (10 $\mu$M), a broad-spectrum tyrosine kinase inhibitor, markedly decreased ACh-induced currents and $Ca^{2+}$ transients, whereas 10 $\mu$M genistin, an inactive analogue, had little effect. Overall these data suggest that the activities of $\alpha3\beta4$ AChRs in MPG neurons are positively regulated by PTK. In conclusion, trosine kinase may be one of the key factors in the regulation of $\alpha3\beta4$ nAChRs in rat MPG neurons, which may play an important roles in the autonomic neuronal function such as synaptic transmission, autonomic reflex, and neuronal plasticity.

• Bulletin of the Korean Chemical Society
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• 제32권12호
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• pp.4352-4360
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• 2011

A four dimensional quantitative structure activity relationship analysis was applied to a series of 50 flavonoid inhibitors of $p56^{lck}$ protein tyrosine kinase by the molecular comparative electron topological method. It was found that the -log (IC50) values of the compounds were highly dependent on the topology, size and electrostatic character of the substituents at seven positions of the flavonoid scaffold in this study. Depending on the negative or positive charge of the groups correctly embedded in these substituents, three-dimensional bio-structure to increase or decrease -log (IC50) values in the training set of 39 compounds was predicted. The test set of 11 compounds was used to evaluate the predictivity of the model. To generate 4D-QSAR model, the defined function groups and pharmacophore used as topological descriptors in the calculation of activity were of sufficient statistical quality ($R^2$ = 0.72 and $Q^2$ = 0.69). Ligand docking approach by using Dock 6.0. These compounds include many flavonoid analogs, They were docked onto human families of p56lck PTKs retrieved from the Protein Data Bank, 1lkl.pdb.

• Bulletin of the Korean Chemical Society
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• 제27권2호
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• pp.266-272
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• 2006

Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were developed for 67 molecules of 2-amino-benzothiazole-6-anilide derivatives against lymphocyte-specific protein tyrosine kinase (P56 LCK). The molecular field analysis (MFA) and receptor surface analysis (RSA) were employed for QSAR studies and the predictive ability of the model was validated by 15 test set molecules. Structure-based investigations using molecular docking simulation were performed with the crystal structure of P56 LCK. Good correlation between predicted fitness scores versus observed activities was demonstrated. The results suggested that the nature of substitutions at the 2-amino and 6-anilide positions were crucial in enhancing the activity, thereby providing new guidelines for the design of novel P56 LCK inhibitors.

• Bulletin of the Korean Chemical Society
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• 제28권2호
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• pp.200-206
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• 2007

Pharmacophore models for lymphocyte-specific protein tyrosine kinase (P56 LCK) were developed using CATALYST HypoGen with a training set comprising of 25 different P56 LCK inhibitors. The best quantitative pharmacophore hypothesis comprises of one hydrogen bond acceptor, one hydrogen bond donor, one hydrophobic aliphatic and one ring aromatic features with correlation coefficient of 0.941, root mean square deviation (RMSD) of 0.933 and cost difference (null cost-total cost) of 66.23. The pharmacophore model was validated by two methods and the validated model was further used to search databases for new compounds with good estimated LCK inhibitory activity. These compounds were evaluated for their binding properties at the active site by molecular docking studies using GOLD software. The compounds with good estimated activity and docking scores were evaluated for physiological properties based on Lipinski's rules. Finally 68 compounds satisfied all the properties required to be a successful inhibitor candidate.

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.756-766
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• 2005

The signaling mechanism underlying aburatubolactam C-induced FasL upregulation was investigated in human Jurkat T cells. After treatment with aburatubolactam C, the src-family PTKs $p56^{lck}\;and\;p59^{fyn}$, and MAP kinases ERK2 and JNK1, were activated prior to FasL upregulation; Both $p56^{lck}\;and\;p59^{fyn}$ were directly activated 2.4- and 2.2-fold, respectively, in vitro by aburatubolactam C. The aburatubolactam C-induced cellular changes, including the activation of ERK2 and INK1, and FasL upregulation, were completely prevented by the PTK inhibitor genistein. The activation of protein kinase C (PKC$\delta,\;\epsilon\;and\;\mu$ was also induced following aburatubolactam C treatment. Although the activation of $p56^{lck}$ and tyrosine phosphorylation of the cellular proteins were not blocked by the PKC inhibitor GFl09203X, the activation of ERK2 was completely abrogated, along with a detectably enhanced JNK1 activation; FasL upregulation, and apoptosis. However, the FasL upregulation and apoptosis were significantly inhibited by the PKC activator PMA, with a remarkable increase in the ERK2 activation. The cytotoxic effect of aburatubolactam C was reduced in the presence of the anti-Fas neutralizing antibody ZB-4. Although ectopic expression of Bcl-2 failed to completely block the cytotoxicity of aburatubolactam C, it was clearly suppressed. The c-Fos mRNA expression was upregulated in a biphasic manner, where the second phasic expression overlapped with the FasL upregulation. Accordingly, these results demonstrate that aburatubolactam C-induced apoptosis is exerted, at least in part, by FasL upregulation dictated by activation of the PTK ($p56^{lck}\;and\;p59^{fyn}$) /JNKI pathway, which is negatively affected by the concurrent activation of the PKC/ERK2 pathway proximal to PTK activation.