• Journal of the Korean Magnetic Resonance Society
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• v.10 no.2
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• pp.163-174
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• 2006

Luteinizing Hormone Releasing Hormone(LHRH) is a decapeptide neurotransmitter known to be regulated by metal ions in the hyperthalamus. Zn-binding LHRH complex was systhesized, and zinc-LHRH complex was studied to understand what kinds of structural modifications would be critical in the LHRH releasing mechanism. Both nonexchangeable and exchangeable $^1H-NMR$ signal assignments were accomplished by pH-dependent and COSY NMR experiments. In addition, $^1H-NMR$ chemical shift changes of a-proton and peptide NH NMR signals at different pH condition, and $^1H-NMR$ signal differences between metal free and metallo-LHRH complex was monitored. NMR signals exhibit that primary metal-binding sites are nitrogens donor of imidazole ring and Arg, and peptide oxygen of Pro-His in the sequence. Structure obtained in this study has a cyclic conformation which is similar to that of energy minimized, and exhibits a specific a-helical turn with residue numbers $(2{\sim}7)$ out of 10 amino acids.

• Journal of the Korean Magnetic Resonance Society
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• v.13 no.2
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• pp.143-153
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• 2009

Luteinizing Hormone Releasing Hormone (LHRH) is composed of 10 amino acids, and is best known as a neurotransmitter. Because of the 80% homology in animals, much more concerns have focused on the substances that have similar functions or can control LHRH. Ni, Cu-LHRH complexes were synthesized. The degree of complexation was monitored by $^1H,\;^{13}C$-NMR chemical shifts, and final products were identified by ESI-Mass spectrum. Solution-state structure determination of Ni-LHRH complex was accomplished by using NMR results and NMR-based distance geometry (DG). Interproton distances from nuclear Overhauser effect spectroscopy (NOESY) were utilized for the molecular structure determination. Results were compared with previous structures obtained from energy minimization and other spectroscopic methods. Structure obtained in this study has a cyclic conformation which is similar to that of energy minimized, and exhibits a specific a-helical turn with residue numbers (2~7) out of 10 amino acids. Comparison of chemical shifts and EPR studies of Ni, Cu-LHRH complexes exhibit that Ni-LHRH complex has same binding sites with the 4-coordination mode as in Zn-LHRH complex.

• Analytical Science and Technology
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• v.13 no.4
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• pp.494-503
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• 2000

A 21-residue peptide corresponding to amino acids 84-104 of $p16^{INK4A}$, the tumor suppressor, has been synthesized and its structure was studied by Circular Dichroism, $^1H$ NMR spectroscopy and molecular modeling. A p16-derived peptide (84-104 amino acids) forming stable complex with CDK4 and CDK6 inhibits the ability of CDK4/6 to phosphorylate pRb in vitro, and blocks cell-cycle progression through G1/S phase as shown in the function of the full-length p16. Its NMR spectral data including NOEs, $^3J_{NH-H{\alpha}}$ coupling constants, $C_{\alpha}H$ chemical shift, the average amplitude of amide chemical shift oscillation and temperature coefficients indicate that the secondary structure of a p16-derived peptide is similar to that of the same region of full-length p16, which consists of helix-turn-helix structure. The 3-D distance geometry structure based on NOE-hased distance and torsion angle restraints is characterized by ${\gamma}$-turn conformation between residues $Gly^{89}-Leu^{91}$(${\varphi}_{i+1}=-79.8^{\circ}$, ${\varphi}_{i+1}=60.2^{\circ}$) as evidenced in a single crystal structure for the corresponding region of p18 or p19, but is undefined at both the N and C termini. This compact and rigid ${\gamma}$-turn region is considered to stabilize the structure of p16-derived peptide and serve as a site recognizing cyelin dependent kinase, and this well-defined ${\gamma}$-turn structure could be utilized for the design of anti-cancer drug candidates.

• Molecules and Cells
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• v.27 no.6
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• pp.673-680
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• 2009

Conversion of the normal soluble form of prion protein, PrP ($PrP^C$), to proteinase K-resistant form ($PrP^{Sc}$) is a common molecular etiology of prion diseases. Proteinase K-resistance is attributed to a drastic conformational change from ${\alpha}$-helix to ${\beta}$-sheet and subsequent fibril formation. Compelling evidence suggests that membranes play a role in the conformational conversion of PrP. However, biophysical mechanisms underlying the conformational changes of PrP and membrane binding are still elusive. Recently, we demonstrated that the putative transmembrane domain (TMD; residues 111-135) of Syrian hamster PrP penetrates into the membrane upon the reduction of the conserved disulfide bond of PrP. To understand the mechanism underlying the membrane insertion of the TMD, here we explored changes in conformation and membrane binding abilities of PrP using wild type and cysteine-free mutant. We show that the reduction of the disulfide bond of PrP removes motional restriction of the TMD, which might, in turn, expose the TMD into solvent. The released TMD then penetrates into the membrane. We suggest that the disulfide bond regulates the membrane binding mode of PrP by controlling the motional freedom of the TMD.

• Proceedings of the Korean Biophysical Society Conference
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• 2003.06a
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• pp.69-69
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• 2003

Human CD99 is a ubiquitous 32-kDa transmembrane protein encoded by the mic2 gene. The major cellular functions of CD99 protein are related to homotypic cell adhension, apoptosis, vesicular protein transport, and differentiation of thymocytes or T cells. Recently it has been reported that expression of a splice variant of CD99 transmembrane protein (Type I and Type II) increases invasive ability of human breast cancer cells. To understand structural basis for cellular functions of CD99 (Type I), we have initiated studies on hCD99$^{TMcytoI}$ and hCD99$^{cytoI}$ using circular dichroism (CD) and multi-dimensional NMR spectroscopy. CD spectrum of hCD99$^{TMcytoI}$ in the presence of 200mM DPC and CHAPS displayed an existence $\alpha$-helical conformation. The solution structure of hCD99$^{cytoI}$ determined by NMR is composed of one N-terminal $\alpha$-helix, $\alpha$A, two C-terminal short $\alpha$-helix segments, $\alpha$B and $\alpha$C. While $\alpha$A and $\alpha$B are connected by the long flexible loop, $\alpha$B and $\alpha$C connected by type III$\beta$-turn. Although it has been rarely figured out the correlation between structure and functional mechanism of hCD99$^{TMcytoI}$ and hCD99$^{cytoI}$, there is possibility of dimerization or oligomerization. In addition, the feasible mechanism of hCD99$^{cytoI}$ is that it could have intramolecular interaction between the N- and C- terminal domain through large flexible AB loop.

• Journal of Microbiology and Biotechnology
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• v.20 no.2
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• pp.301-310
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• 2010

Bacillus subtilis strains produce a broad spectrum of bioactive peptides. The lipopeptide surfactin belongs to one well-known class, which includes amphiphilic membrane-active biosurfactants and peptide antibiotics. Both the srfA promoter and the ComP-ComA signal transduction system are an important part of the factor that results in the production of surfactin. Bs-M49, obtained by means of low-energy ion implantation in wild-type Bs-916, produced significantly lower levels of surfactin, and had no obvious effects against R. solani. Occasionally, we found strain Bs-M49 decreased spore formation and the development of competence. Blast comparison of the sequences from Bs-916 and M49 indicate that there is no difference in the srfA operon promoter PsrfA, but there are differences in the coding sequence of the comA gene. These differences result in three missense mutations within the M49 ComA protein. RT-PCR analyses results showed that the expression levels of selected genes involved in competence and sporulation in both the wild-type Bs-916 and mutant M49 strains were significantly different. When we integrated the comA ORF into the chromosome of M49 at the amyE locus, M49 restored hemolytic activity and antifungal activity. Then, HPLC analyses results also showed the comA-complemented strain had a similar ability to produce surf actin with wild-type strain Bs-916. These data suggested that the mutation of three key amino acids in ComA greatly affected the biological activity of Bacillus subtilis. ComA protein 3D structure prediction and motif search prediction indicated that ComA has two obvious motifs common to response regulator proteins, which are the N-terminal response regulator receiver motif and the C-terminal helix-turn-helix motif. The three residues in the ComA N-terminal portion may be involved in phosphorylation activation mechanism. These structural prediction results implicate that three mutated residues in the ComA protein may play an important role in the formation of a salt-bridge to the phosphoryl group keeping active conformation to subsequent regulation of the expression of downstream genes.

• Food Science and Biotechnology
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• v.18 no.1
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• pp.228-233
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• 2009

This study was conducted to examine the changes in the molecular structure and physiological activities of silk fibroin by gamma irradiation. The results of gel permeation chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the molecular weight of fibroin was increased depending upon the irradiation dose. Secondary structure of fibroin determined by using circular dichroism revealed that the ratio of $\alpha$-helix was increased up to 10 kGy and then decreased depending upon the irradiation dose. Whereas, the ratio of $\beta$-sheet, $\beta$-turn, and random coil were decreased and then increased with an alteration in the $\alpha$-helix secondary conformation. The 2.2-diphenyl-1-picryl-hydrazil (DPPH) radical scavenging activity of fibroin was increased by gamma irradiation at 5 kGy, but was decreased above 10 kGy depending upon the irradiation dose. Also, the inhibition activities of tyrosinase and melanin synthesis of fibroin were increased by gamma irradiation. These results indicated that gamma irradiation could be used as an efficient method to make fibroin more suitable for the development of functional foods and cosmetics.