Chul-Hak Yang

Distinct Roles of the N-terminal-binding Domain and the C-terminal-solubilizing Domain of α-Synuclein, a Molecular Chaperone

α-Synuclein, an acidic neuronal protein of 140 amino acids, is extremely heat-resistant and is natively unfolded. Recent studies have demonstrated that α-synuclein has chaperone activity both in vitro and in vivo, and that this activity is lost upon removing its C-terminal acidic tail. However, the detailed mechanism of the chaperone action of α-synuclein remains unknown. In this study, we investigated the molecular mechanism of the chaperone action of α-synuclein by analyzing the roles of its N-terminal and C-terminal domains. The N-terminal domain (residues 1–95) was found to bind to substrate proteins to form high molecular weight complexes, whereas the C-terminal acidic tail (residues 96–140) appears to be primarily involved in solubilizing the high molecular weight complexes. Because the substrate-binding domain and the solubilizing domain for chaperone function are well separated in α-synuclein, the N-terminal-binding domain can be substituted by other proteins or peptides. Interestingly, the resultant engineered chaperone proteins appeared to display differential efficiency and specificity in terms of the chaperone function, which depended upon the nature of the binding domain. This finding implies that the C-terminal acidic tail of α-synuclein can be fused with other proteins or peptides to engineer synthetic chaperones for specific purposes.

Inhibition of fos–jun–DNA complex formation by dihydroguaiaretic acid and in vitro cytotoxic effects on cancer cells

The effect of dihydroguaiaretic acid (DHGA), isolated from the aryls of Myristica fragrans, on the transcription factor (fos-jun dimer) action was investigated via in vitro assay. DHGA showed an inhibitory effect against the complex formation of the fos-jun dimer and the DNA consensus sequence with an IC50 value of 0.21 micromol. Nordihydroguaiaretic acid (NDGA) and curcumin also inhibited fos-jun dimer action showing IC50 values of 7.9 and 6.9 nmol, respectively. DHGA and NDGA suppressed leukemia, lung cancer and colon cancer in an in vitro bioassay. The in vitro experiment suggested that inhibition of fos-jun-DNA complex formation could be due to the direct interference of fos-jun dimer binding onto the DNA consensus sequence by NDGA and curcumin.

New and known symmetrical curcumin derivatives inhibit the formation of Fos-Jun-DNA complex

We previously reported that curcumin, the yellow pigment of turmeric, inhibited the formation of the Fos-Jun-DNA complex. Thus, we have synthesized 12 symmetrical curcuminoids. We used a slightly modified version of Pabons method to search for an inhibitor more potent than curcumin. Of the synthesized curcuminoids, BJC005, CHC011, and CHC007 exhibited a remarkably high inhibitory activity. Their IC(50) values are 5.4 microM, 0.30 mM, and 0.38 mM, respectively. These IC(50) data indicated that BJC005 is nearly 90 times more effective than curcumin. The BJC005 has shown a more powerful profile than momordin, which, until now, has been reported as a potent Fos-Jun inhibitor. Also BJC005 and CHC007 have not been synthesized before. We report for the first time that the novel BJC005 and CHC007 exhibit highly inhibitory activity against transcription activity.

Analysis of FcγRIII and IgG Fc Polymorphism Reveals Functional and Evolutionary Implications of Protein–Protein Interaction

Abstract. Fcγ receptor III (FcγRIII), a low-affinity receptor for the Fc portion of immunoglobulin G (IgG Fc), targets antigen-antibody complexes in a variety of effector cells of the immune system. We have investigated FcγRIII and IgG Fc polymorphism and made comparative analysis of the functional and evolutionary implications of the interaction between these two molecules. Sequence analysis and comparison of the three-dimensional structure suggest that the C-terminal Ig domain of FcγRIII is associated with the binding of IgG. The polymorphic residues of FcγRIII are mainly located in the region of the C-terminal Ig domain that might be involved in IgG binding. Therefore, polymorphism and functional binding affinity seems to be related to each other as has been increasingly implicated in clinical observations. IgG Fcs, the natural ligand of FcγRs, also exhibit significant polymorphism. Three regions have been identified where polymorphism frequently occurs: the putative FcR binding site, the linker region, and the intermolecular domain-domain interface of the second Ig domain. The putative FcγR binding sites where polymorphic, and isotype-specific residues cluster are consistent with the regions that have been identified by mutagenesis and molecular modeling studies. The polymorphic residues of IgG Fc were mainly located in the molecular surface, which could be used in the recognition of other binding molecules. These observations suggest that polymorphic and isotype-specific residues in IgG Fc are closely related to their function and protein-protein interaction. Therefore, the colocalization of the polymorphic residues of FcγRIII and IgG Fcs at their docking sites implies that the polymorphic residues would affect the IgG-FcγRIII binding interactions to optimize their signaling through evolution.

Molecular dynamics simulations of cyclohenicosakis-[(1→2)-β-d-gluco-henicosapyranosyl], a cyclic (1→2)-β-d-glucan (a ‘cyclosophoraose’) of DP 21

Abstract We report molecular dynamics simulations of cyclohenicosakis-[(1→2)-β- d - gluco -henicosapyranosyl], termed ‘cyclosophohenicosamer’, a member of a class of cyclic (1→2)-β- d -glucans (‘cyclosophoraoses’). Our goals were to provide insights into the conformational preferences of these cyclosophoraoses. Simulated annealing and constant-temperature molecular dynamics calculations were performed on the DP 21 cyclosophohenicosamer. The radius of gyration ( R G ) of the molecule and the conformation of glycosidic dihedral angles were used to analyze the result of computational studies. Most glycosidic linkages were concentrated in the lowest-energy region of the φ – ψ energy map, and the values of radius of gyration from our simulations were consistent with the reported experimental value. The simulations produced various types of compact and asymmetric conformations within reasonable ranges of the glycosidic linkage conformation and radius of gyration. The results indicate the presence of a high degree of molecular flexibility of cyclosophohenicosamer and suggest the uniqueness of inclusion complexation with other molecules through this molecular flexibility.

Quad-Band Antenna With High Isolation MIMO and Broadband SCS for Broadcasting and Telecommunication Services

The quad-band antenna, which is composed of two compact dual-band antennas, is presented for portable media player (PMP) applications. The antenna for the broadcasting dual band (DVB-H UHF: 470-862 MHz; L: 1452-1492 MHz) is composed of a planar inverted-F antenna (PIFA) with self-complementary structure (SCS) for the broadband performance. The measured operation bandwidth of the antenna (VSWR <; 4) is 500 MHz (370-870 MHz) at UHF band and 220 MHz (1300-1520 MHz) at L band. The two-channel WLAN multiple-input-multiple-output (MIMO) antenna with high isolation performance for the telecommunication dual band (WLAN 11b: 2.4-2.5 GHz; 11a: 5.15-5.825 GHz) is composed of a PIFA operating at 2-GHz band and a loop antenna operating at 5-GHz band. The proposed antennas are fabricated in a PMP case (εr = 3.2 ).

Inhibition of AP‐1 transcription activator induces myc‐dependent apoptosis in HL60 cells

Transcriptional activation of AP‐1 is intricately involved in cell proliferation and transformation. The natural product, nordihydroguaiaretic acid (NDGA) shows an inhibitory effect on the binding of jun/AP‐1 protein to the AP‐1 site in 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA)‐stimulated HL60 cells. The NDGA inhibits the auto‐regulated de novo synthesis of c‐jun mRNA in TPA‐stimulated HL60 cells. Our data also determine that this compound induces proliferation inhibition and apoptosis in human leukemia HL60 cells. To obtain information on the functional role of the AP‐1 inhibition by NDGA in apoptosis signaling, the effects of pharmacological inhibition of AP‐1 binding on c‐myc, p53, and bax protein level were determined. Our results indicate that treatment of cells with NDGA enhances c‐myc, p53, and bax protein levels. To rule out the possibility that NDGA will induce apoptosis because of the effects on proteins other than AP‐1, we investigated the effect of another AP‐1 inhibitor, SP600125, which is specific to Jun‐N‐terminal kinase. SP600125 decreased not only the phosphorylation level of jun protein but also AP‐1/DNA binding activity. Also, apoptosis was observed to be induced by SP600125, concomitant with the increase in c‐myc, p53, and bax protein level. In addition, apoptosis induced by both AP‐1 inhibitors was accompanied by the activation of a downstream apoptotic cascade such as caspase 9, caspase 3, and poly[ADP‐ribose]polymerase (PARP). When the cells were treated with NDGA or SP600125 in the presence of antisense c‐myc oligonucleotides, apoptosis was not observed and an increase of c‐myc, p53, and bax proteins was not manifested. All these results show that the inhibition of the transcription factor AP‐1 action is related with either the drug‐induced apoptosis or the drug toxicity of the HL60 cells. The apoptosis induced by AP‐1 inhibition may be dependent on c‐myc protein levels suggesting that the c‐myc protein induces apoptosis at a low level of AP‐1 binding activity. Altogether, our findings suggest that the presence of the AP‐1 signal acts as a survival factor that determines the outcome of myc‐induced proliferation or apoptosis.

Monte Carlo simulations of the chiral recognition of fenoprofen enantiomers by cyclomaltoheptaose (β-cyclodextrin)

Abstract Differential complexation of fenoprofen enantiomers by cyclomaltoheptaose (β-cyclodextrin) was investigated by Monte Carlo docking simulations. The chiral discrimination of ( R )- and ( S )-fenoprofen by β-cyclodextrin was discussed in terms of the difference in the interaction energies and the patterns of molecular interactions. The interaction energies between each enantiomer of fenoprofen and β-cyclodextrin were consistent with the reported experimental results that showed that the S isomer interacted preferentially with β-cyclodextrin and was retained longer in a separation process than the R isomer. The thermodynamic preference of inclusion complex formation of ( S )-fenoprofen could be explained by the orientation of the phenyl group attached to the chiral carbon, which provided closer contact and thus more favorable intermolecular interactions between the host and guest molecule. The results presented here would be very useful for the prediction of chiral recognition ability of β-cyclodextrin.

Crystallization and preliminary X-ray studies of hORF6, a novel human antioxidant enzyme

HORF6 is a member of the novel antioxidant enzyme family found in humans. A recombinant form of hORF6 expressed and purified from E. coli has been crystallized by the hanging-drop method using various PEGs as precipitating agents. HORF6 crystallizes in two different monoclinic space groups, P21 and C2. The P21 crystals have unit-cell dimensions of a = 47.85, b = 75.17, c = 63.30 A and beta = 110.21 degrees and contain two monomers per asymmetric unit, while the C2 crystals have unit-cell dimensions of a = 165.27, b = 95.44, c = 166.44 A and beta = 128.97 degrees and contain more than six monomers per asymmetric unit. The P21 crystals with the smaller unit cell diffract X-rays better and behave well for the X-ray analysis. A native data set from a single crystal of the P21 space group gas been collected to 2.0 A resolution.

Unsaturated fatty acids bind Myc–Max transcription factor and inhibit Myc–Max–DNA complex formation

Oncoprotein Myc, hetero-dimerized with Max through a b/HLH/Zip region, is a transcription factor that governs important cellular processes such as cell cycle entry, proliferation and differentiation. We found that linoleic acid, isolated from Pollen Typhae, and other unsaturated fatty acids have strong inhibitory effects on the binding of Myc-Max heterodimer to an E-box DNA site (CA(C/T)GTG). The interaction of a fatty acid with a protein dimer, not with DNA, is assumed to block the entire Myc-Max-DNA complex formation. Unsaturated fatty acids also showed cytotoxicity against a SNU16 human stomach cancer cell line and conjugated linoleic acid suppressed mRNA expression of several myc-target genes; ornithine decarboxylase, p53, cdc25a in the SNU16 cells.