• Bulletin of the Korean Chemical Society
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• v.30 no.3
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• pp.623-629
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• 2009

We have performed replica-exchange molecular dynamics simulations on 41 residue peptide mainly composed of NAC (non A$\beta$ component) sequence in $\alpha$-Synuclein. To investigate conformational characteristics of intrinsically unstructured peptides, we carried out structural analysis on the ‘representative structures’ for ensemble of structures occurring at different temperatures. The secondary structure profile obtained from our simulations suggests that the NAC region of $\alpha$-synuclein can be divided into roughly three helical-like segments. It is found that the overall helix-turn-helix like topology is conserved even though the conformational fluctuations grow as the temperature increases. The coordinate-based and the distance-based representative structures exhibit noticeable differences at higher temperatures while they are similar at lower temperatures. It is found that structural variations for the coordinate-based representative structures are much larger, suggesting that distance-based representative structures provide more reliable information concerning characteristic features of intrinsically unstructured proteins. The present analysis also indicates that the conformational features of representative structures at high temperatures might be related to those in membrane or low pH environment.

• Proceedings of the Korean Magnetic Resonance Society Conference
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• 2002.08a
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• pp.35-35
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• 2002

• Journal of the Korean Magnetic Resonance Society
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• v.2 no.1
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• pp.33-40
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• 1998

The cross peak intensities versus mixing times of 2D NOESY spectrum for a corepressor L-trp were simulated for the case of a ligand exchanging between free (AX) and bound (A'X') forms in protein/DNA complex. The direct NOE (I(AX)) of the free ligand exhibited a small positive intensity indicative of the strong dominant influence of the bound ligand. The exchange-mediated NOE peak (I(AX')) was very sensitive to corepressor exchange. However, both diagonal (I(A'A')) and direct NOE (I(A'X')) intensities of the bound ligand were not affected much at initial stage. Both peaks were severely influenced by exchange at mixing times of greater than 100 ms. In conclusion, since the NOE intensity is a function of exchange rate, the exchange effect should be considered to properly extract accurate distance information for bound ligand in the presence of conformational exchange.

• Journal of the Korean Magnetic Resonance Society
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• v.14 no.2
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• pp.117-126
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• 2010

Hsp33 is a molecular chaperone achieving a holdase activity upon response to a dual stress by heat and oxidation. Despite several crystal structures available, the activation process is not clearly understood, because the structure inactive Hsp33 as its reduced, zinc-bound, monomeric form has not been solved yet. Thus, we initiated structural investigation of the reduced Hsp33 monomer by NMR. In this study, to overcome the high molecular weight (33 kDa), the protein was triply isotope-[$^{13}C$, $^{15}N$, $^2H$]-labeled and its inactive, monomeric state was ensured. 2D-[$^1H$, $^{15}N$]-TROSY and a series of triple resonance spectra could be successfully obtained on a high-field (900 MHz) NMR machine with a cryoprobe. However, under all of the different conditions tested, the number of resonances observed was significantly less than that expected from the amino acid sequence. Thus, a possible contribution of dynamic conformational exchange leading to a line broadening is suggested that might be important for activation process of Hsp33.

• Journal of the Korean Magnetic Resonance Society
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• v.6 no.2
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• pp.94-102
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• 2002

An interesting recent application of intermolecular NOE experiment is the saturation transfer difference NMR(STD-NMR) method that is useful in screening compound libraries to identify bio-active ligands. This technique also identifies the group epitopes of the bound ligand in a reversibly forming protein-ligand complex. We present here a complete relaxation and conformational exchange matrix (CORCEMA) theory (Moseley et al., J. Magn. Reson. B, 108, 243-261 (1995)) applicable for the STD-NMR experiment. Using some ideal model systems we have analyzed the factors that influence the STD intensity changes in the ligand proton NMR spectrum when the resonances from some protons on the receptor protein are saturated. These factors will be discussed and some examples of its application in some model systems will be presented. This CORCEMA theory for STD-NMR and the associated algorithm are useful in a quantitative interpretation of the STD-NMR effects, and are likely to be useful in structure-based drug design efforts. They are also useful in a quantitative characterization of protein-protein (or protein-nucleic acid) contact surfaces from an intermolecular cross-saturation NMR experiment.

• Bulletin of the Korean Chemical Society
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• v.34 no.5
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• pp.1401-1406
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• 2013

This study tested the feasibility of observing H/D exchange of intact protein by top-down electron capture dissociation (ECD) mass spectrometry for the investigation of protein structure. Ubiquitin is selected as a model system. Local structural information was obtained from the deuteration levels of c and $z^{\cdot}$ ions generated from ECD. Our results showed that ${\alpha}$-helix region has the lowest deuteration level and the C-terminal fraction containing a highly mobile tail has the highest deuteration level, which correlates well with previous X-Ray and HDX/NMR analyses. We studied site-specific H/D exchange kinetics by monitoring H/D exchange rate of several structural motives of ubiquitin. Two hydrogen bonded ${\beta}$-strands showed similar HDX rates. However, the outer ${\beta}$-strand always has higher deuteration level than the inner ${\beta}$-strand. The HDX rate of the turn structure (residues 8-11) is lower than that of ${\beta}$-strands (residues 1-7 and residues 12-17) it connects. Although isotopic distribution gets broader after H/D exchange which results in a limited number of backbone cleavage sites detected, our results demonstrate that this method can provide valuable detailed structural information of proteins. This approach should also be suitable for the structural investigation of other unknown proteins, protein conformational changes, as well as protein-protein interactions and dynamics.

• Bulletin of the Korean Chemical Society
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• v.25 no.5
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• pp.609-616
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• 2004

Complexation of cesium ion by p-tert-butylcalix[6]arene ethyl ester was studied by NMR spectroscopy in nonpolar $CDCl_3$ and polar acetone-$d_6$ and the results were compared with each other. Analysis of temperature dependent $^1H$ spectra and titration curves reveals that both solvents result in a 1 : 1 cone-form complex with nonpolar $CDCl_3$yielding a more tightly bound one than acetone-$d_6$. Unexpectedly, at very low temperature, we have found that two phenyl ring proton peaks of equal intensity appear both in $CDCl_3$and in acetone-$d_6$ solution which gradually collapse and eventually coalesce into a single line as temperature is raised. This observation could be interpreted in terms of the chemical exchange through direct and/or indirect interconversion between two equivalent conformations possible the complex in both solvents over the temperature range observed. And broadening of $^{133}Cs$ (I = 7/2) nmr line with increasing temperature has also been observed, indicating the exchange of $^{133}Cs$ ion between the complex and the solvent. From numerical fitting of lineshape changes for one-dimensional $^1H$ and $^{133}Cs$ spectra, the exchange rate constants and other relevant parameters for this conformational interconversion and the complex-solvent exchange were deduced.

• Proceedings of the Korean Biophysical Society Conference
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• 1996.07a
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• pp.27-27
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• 1996

SecA protein which has a pivotal role in the preprotein cranslocation across the inner membrane of Escherichia coli is a water-soluble protein with an unusual property of penetrating the membrane readily. An interesting and important question is what structural characteristics of SecA enables its ready penetration of lipid bilayer. The conformational properties of SecA in solution at 3$0^{\circ}C$, pH 7.5 were observed by hydrogen-tritium (HT) exchange, and denaturant-induced denaturation/renaturation and thermal unfolding. (omitted)

• Bulletin of the Korean Chemical Society
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• v.18 no.9
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• pp.961-965
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• 1997

1H NMR spectra of 3,3-dimethylpiperidine (1) at -70 to 30 ℃ exhibit gradual change from slow to rapid exchange between two alternate chair forms. The exchange rate constant was determined as a function of temperature by simulating the line shape of the signal from the two methyl groups using the modified Bloch equations. The resulting free energy of activation is ΔG* = 44.4±1.9 kJ mol-1 at 298 K. The 1H NMR spectrum of a D2O or dimethylsulfoxide-d6 (DMSO-d6) solution containing 1 and [SiW11CoⅡO39]6- exhibits separate signals for the free ligand and the complex, indicating that the ligand exchange is slow on the NMR time scale. In D2O the piperidine ring is frozen as a chair form even at room temperature with the cobalt ion bonded to the axial position of the nitrogen atom. When DMSO-d6 is added to the D2O solution, the NMR spectral change suggests that a rapid exchange occurs between the chair form and another conformer. It is proposed that the conformation of ^b1^b coordinated to [SiW11CoⅡO39]6- in DMSO-d6 is close to a twist form.

• Biomolecules & Therapeutics
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• v.29 no.5
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• pp.527-535
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• 2021

Sclerostin (SOST), a regulator of bone formation in osteocytes, inhibits the canonical Wnt signaling by interacting with low-density lipoprotein receptor-related protein 5/6 (LRP5/6) to prevent Wnt binding. Loss-of-function mutations of the SOST gene caused massive bone outgrowth and SOST-null mouse exhibited a high bone density phenotype. Therefore, SOST has been suggested as a promising therapeutic target for osteoporosis. A few previous studies with X-ray crystallography identified the binding interfaces between LRP6 and SOST, but there are limitations in these studies as they used truncated SOST protein or SOST peptide. Here, we analyzed the conformational dynamics of SOST-LRP6 E1E2 complex using hydrogen/deuterium exchange mass spectrometry (HDX-MS). We examined the effect of the C-terminal tail of SOST on LRP6 conformation upon complex formation. HDX-MS analysis suggested a new potential binding interface for the C-terminal region of SOST that was missing from the previous crystal structure of the SOST-LRP6 E1E2 complex.