• Biodesign
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• v.6 no.4
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• pp.92-95
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• 2018

API5 is a unique oncogenic, non-BIR type IAP nuclear protein and is up-regulated in several cancers. It exerts several functions, such as apoptosis inhibition, cell cycle progression, cancer immune escape, and anticancer drug resistance. Although structural studies of API have revealed that API5 mediates protein-protein interactions, its detailed molecular functions remain unknown. Since FGF2 is one of API5's major interacting proteins, structural studies of the API5-FGF2 complex will provide insight into both proteins' molecular function. We overexpressed and purified API5 and FGF2 in Escherichia coli and crystallized the API-FGF2 complex using polyethylene glycol (PEG) 6000 as a precipitant. Diffraction data were collected to a $2.7{\AA}$ resolution using synchrotron X-rays. Preliminary diffraction analysis revealed that the API5-FGF2 complex crystal belongs to the space group $P2_12_12_1$ with the following unit cell parameters: a = 46.862, b = 76.523, $c=208.161{\AA}$. One asymmetric unit with 49.9% solvent contains one API5-FGF2 complex. Molecular replacement calculation, using API5 and FGF2 coordinates, provided a clear electron density map for an API5-FGF2 complex.

• Journal of the Korean Magnetic Resonance Society
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• v.27 no.3
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• pp.17-22
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• 2023

Bloom syndrome protein (BLM) is a pivotal RecQ helicase necessary for genetic stability through DNA repair processes. Our investigation focuses on the N-terminal region of BLM, which has been considered as an intrinsically disordered region (IDR). This IDR plays a critical role in DNA metabolism by interacting with other proteins. In this study, we performed triple resonance experiments of BLM_220-300 and presented the backbone chemical shifts. The secondary structure prediction based on chemical shifts of the backbone atoms shows the region is disordered. Our data could help further interaction studies between BLM_220-300 and its binding partners using NMR.

• BMB Reports
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• v.34 no.1
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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.

• Proceedings of the SCSK Conference
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• 2003.09a
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• pp.590-600
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• 2003

The matrix metalloproteinases (MMPs) are a family of enzymes responsible for degrading connective tissue. MMPs catalyze the breakdown of collagen from the extracellular matrix, leading to wrinkle formation and accelerated skin aging. Furthermore, ultraviolet irradiation causes increased expression of certain MMPs. In the extracellular matrix turnover, MMPs are interacting with endogenous regulators named tissue inhibitors of metalloproteinases (TIMPs). Using peptide substrate assays, it has been demonstrated that TIMP-MMP complexes interact highly specifically with $K_{i}$ values of 10$^{-9}$ -10$^{-16}$ M. Therefore applications for TIMP as inhibitor of collagen degradation are suggested for cosmetic anti-aging products to prevent wrinkle formation and loss of elasticity. To date four TIMP proteins (TIMP-1, TIMP-2, TIMP-3 and TIMP-4) have been identified which show a high degree in sequence similarity. The production of human TIMP-2, a 194-residue nonglycosylated protein, was performed by fed-batch culture of Escherichia coli. TIMP-2 accumulated in the bacterial cells in an insoluble form as inclusion bodies. The inclusion bodies were solubilized and the protein refolded to yield the native TIMP-2 in the active form. The integrity of the protein was confirmed by mass analysis, Edman sequencing and gel shift experiments with authentic samples. The inhibitory activity of the refolded and purified TIMP-2 was demonstrated with MMP-1 and MMP-2 assays using synthetic fluorogenic peptide substrates.s.

• BMB Reports
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• v.31 no.6
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• pp.611-614
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• 1998

Protein kinase CKII (CKII) is a protein Ser/Thr kinase that is ubiquitously distributed in eukaryotic cells. Although it has been suggested that CKII plays an critical role in cell growth and proliferation, its functional significance and regulation in the cells remain poorly understood. To investigate the exact biological function of CKII, we have identified proteins that interact with the subunits of CKII using the twohybrid system. In this report, we have identified cathepsin L, a lysosomal protease, as a cellular protein capable of interacting with the ${\beta}$ subunit of CKII. Cathepsin L does not interact with the ${\alpha}$ subunit of CKII, supporting the idea that the ${\beta}$ subunit can mediate the interaction of CKII with target proteins. We have found that cathepsin L has several putative CKII phosphorylation sites including Thr-84, Ser-160, Ser-270, Thr-288, and Ser-301. These data suggest that CKII is a possible protein kinase for cathepsin L phosphorylation.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.2
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• pp.95-100
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• 2011

DREAM (downstream regulatory element antagonistic modulator) is a calcium-binding protein that regulates dynorphin expression, promotes potassium channel surface expression, and enhances presenilin processing in an expression level-dependent manner. However, no molecular mechanism has yet explained how protein levels of DREAM are regulated. Here we identified group I mGluR (mGluR1/5) as a positive regulator of DREAM protein expression. Overexpression of mGluR1/5 increased the cellular level of DREAM. Up-regulation of DREAM resulted in increased DREAM protein in both the nucleus and cytoplasm, where the protein acts as a transcriptional repressor and a modulator of its interacting proteins, respectively. DHPG (3,5-dihydroxyphenylglycine), a group I mGluR agonist, also up-regulated DREAM expression in cortical neurons. These results suggest that group I mGluR is the first identified receptor that may regulate DREAM activity in neurons.

• Development and Reproduction
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• v.22 no.1
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• pp.65-72
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• 2018

Cyclic AMP-response element binding protein zhangfei (CREBZF), a member of ATF/CREB (activating transcription factor/ cAMP response element binding protein) family, regulates numerous cellular functions and development of cells by interacting transcription factors. This study discovered the expression pattern of CREBZF in seminiferous tubule of testes during the postnatal development of mice. In testis, CREBZF mRNA expression was the highest among other organs. Immunofluorescence analyses showed that the CREBZF was specifically expressed on spermatocyte but not in spermatogonia and Sertoli cells in seminiferous epithelium of mouse testis. Semi-quantitative polymerase chain reaction (PCR) analysis showed that CREBZF transcript level was significantly elevated during postnatal development of mouse testis. Confocal imaging analysis indicated that the protein expression of CREBZF in seminiferous tubule remained low until postnatal day (PD) 14, and was dramatically increased in PD 21. Interestingly, only one type of the spermatocyte expressed CREBZF specifically among SCP3-positive spermatocytes. Taken together, these results suggest that CREBZF may be novel putative marker of the spermatocyte and regulate meiosis during postnatal development of mice.

• Journal of Life Science
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• v.28 no.5
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• pp.597-604
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• 2018

Although the core mechanisms of Attention Deficit/Hyperactivity Disorder (ADHD) are unknown, several ADHD-associated proteins have been studied. G-protein - coupled receptor kinase interacting protein-1 (GIT1) is a multifunctional adapter protein that affects neuron growth and dendrite formation. GIT1-deficient mice have shown ADHD-like behavior and also recovered through amphetamine treatment. In this study, gliotransmitters were investigated in both intracellular and extracellular space from GIT1-deficient mice. To measure the amount of gliotransmitters, primary astrocyte cultures were taken from the cerebral and cerebellar cortices of wild (WT), hetero (HE), and knock-out (KO) mice. Major gliotransmitters were analyzed using high-performance liquid chromatography. It was observed that the amount of excitatory and inhibitory gliotransmitters were dependent on genotype and showed a change in excitation/inhibition ratios. Interestingly, the major excitatory gliotransmitter, glutamate, existed at the lowest level in WT mice, but the amount of inhibitory gliotransmitters, gamma-aminobutyric acid (GABA) and glycine, varied depending on brain region. Remarkably, an increased amount of GABA was measured at the intracellular cerebrum in WT mice compared with KO mice. It was presumed that KO mice would secrete more inhibitory gliotransmitters to compensate for GIT1 depletion or else acquire a defect to reuptake-secreted GABA. This may be a possible mechanism for ADHD pathology.

• Proceedings of the Korean Society of Sericultural Science Conference
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• 2004.04a
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• pp.38-38
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• 2004

• Proceedings of the PSK Conference
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• 2003.04a
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• pp.254.2-254.2
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• 2003

For many viruses, -1 ribosomal frameshifting regulate protein synthesis using an RNA pseudoknot. The integrity of pseudoknot stability and structure is the important feature for efficient frameshifting. Thus, small molecules interacting with viral RNA pseudoknots would be potential antiviral agents targeting\ulcorner frameshifting system in viruses. X-ray structure of RNA pseudoknot complexed with biotin has been reported, in which biotin is bound at the interface between the pseudoknot's stacked helices. (omitted)