Sunggoo Yun

An Electrochemically Controllable Nanomechanical Molecular System Utilizing Edge-to-Face and Face-to-Face Aromatic Interactions

[formula: see text] A new molecular system, 2,11-dithio[4,4]metametaquinocyclophane containing a quinone moiety, was designed and synthesized. As the quinone moiety can readily be converted into an aromatic pi-system (hydroquinone) upon reduction, the nanomechanical molecular cyclophane system exhibits a large flapping motion like a molecular flipper from the electrochemical redox process. The conformational changes upon reduction and oxidation are caused by changes of nonbonding interaction forces (devoid of bond formation/breaking) from the edge-to-face to face-to-face aromatic interactions and vice versa, respectively.

Roles of dimerization in folding and stability of ketosteroid isomerase from Pseudomonas putida biotype B

Equilibrium and kinetic analyses have been performed to elucidate the roles of dimerization in folding and stability of KSI from Pseudomonas putida biotype B. Folding was reversible in secondary and tertiary structures as well as in activity. Equilibrium unfolding transition, as monitored by fluorescence and ellipticity measurements, could be modeled by a two‐state mechanism without thermodynamically stable intermediates. Consistent with the two‐state model, one dimensional (1D) NMR spectra and gel‐filtration chromatography analysis did not show any evidence for a folded monomeric intermediate. Interestingly enough, Cys 81 located at the dimeric interface was modified by DTNB before unfolding. This inconsistent result might be explained by increased dynamic motion of the interface residues in the presence of urea to expose Cys 81 more frequently without the dimer dissociation. The refolding process, as monitored by fluorescence change, could best be described by five kinetic phases, in which the second phase was a bimolecular step. Because <30% of the total fluorescence change occurred during the first step, most of the native tertiary structure may be driven to form by the bimolecular step. During the refolding process, negative ellipticity at 225 nm increased very fast within 80 msec to account for >80% of the total amplitude. This result suggests that the protein folds into a monomer containing most of the α‐helical structures before dimerization. Monitoring the enzyme activity during the refolding process could estimate the activity of the monomer that is not fully active. Together, these results stress the importance of dimerization in the formation and maintenance of the functional native tertiary structure.

Carceroisomerism and twistomerism in C4c tetraoxatetrathiahemicarceplexes

Four new C4v tetraoxatetrathiahemicarceplexes were synthesized and characterized. Their carceroisomers and half-twistomers were simultaneously observed by 1H NMR spectra at low temperature. The largest isomerization energy barrier of carceroisomers was 15.5 kcal mol−1 and the isomerization energy barriers of twistomers are significantly larger than those of carceroisomers.

Trifluoroethanol Increases the Stability of Δ5-3-Ketosteroid Isomerase 15N NMR RELAXATION STUDIES

In the equilibrium unfolding process of Δ5-3-ketosteroid isomerase from Pseudomonas testosteroni by urea, it was observed that the enzyme stability increases by 2.5 kcal/mol in the presence of 5% trifluoroethanol (TFE). To elucidate the increased enzyme stability by TFE, the backbone dynamics of Δ5-3-ketosteroid isomerase were studied in the presence and absence of 5% TFE by 15N NMR relaxation measurements, and the motional parameters (S 2, τ e , and R ex) were extracted from the relaxation data using the model-free formalism. The presence of 5% TFE causes little change or a slight increase in the order parameters (S 2) for a number of residues, which are located mainly in the dimer interface region. However, the majority of the residues exhibit reduced order parameters in the presence of 5% TFE, indicating that high frequency (pico- to nanosecond) motions are generally enhanced by TFE. The results suggest that the entropy can be an important factor for the enzyme stability, and the increase in entropy by TFE is partially responsible for the increased stability of Δ5-3-ketosteroid isomerase.