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Contribution of a Low-Barrier Hydrogen Bond to Catalysis Is Not Significant in Ketosteroid Isomerase
장도수,최길돈,차형진,신세정,홍비학,이희천,이형주,최관용 한국분자세포생물학회 2015 Molecules and cells Vol.38 No.5
Low-barrier hydrogen bonds (LBHBs) have been proposed to have important influences on the enormous reaction rate increases achieved by many enzymes. Δ5-3-ketosteroid isomerase (KSI) catalyzes the allylic isomerization of Δ5-3- ketosteroid to its conjugated Δ4-isomers at a rate that approaches the diffusion limit. Tyr14, a catalytic residue of KSI, has been hypothesized to form an LBHB with the oxyanion of a dienolate steroid intermediate generated during the catalysis. The unusual chemical shift of a proton at 16.8 ppm in the nuclear magnetic resonance spectrum has been attributed to an LBHB between Tyr14 Oη and C3-O of equilenin, an intermediate analogue, in the active site of D38N KSI. This shift in the spectrum was not observed in Y30F/Y55F/D38N and Y30F/Y55F/Y115F/D38N mutant KSIs when each mutant was complexed with equilenin, suggesting that Tyr14 could not form LBHB with the intermediate analogue in these mutant KSIs. The crystal structure of Y30F/Y55F/Y115F/D38N-equilenin complex revealed that the distance between Tyr14 Oη and C3-O of the bound steroid was within a direct hydrogen bond. The conversion of LBHB to an ordinary hydrogen bond in the mutant KSI reduced the binding affinity for the steroid inhibitors by a factor of 8.1-11. In addition, the absence of LBHB reduced the catalytic activity by only a factor of 1.7-2. These results suggest that the amount of stabilization energy of the reaction intermediate provided by LBHB is small compared with that provided by an ordinary hydrogen bond in KSI.
차형진,장도수,김연길,최관용 한국구조생물학회 2015 Biodesign Vol.3 No.2
H-bond networks that are formed by polar or charged residues play a critical role in the structure, catalytic activity andstability of proteins. Ketosteroid isomerase (KSI) catalyzes the allylic isomerization of a variety of Δ5-3-ketosteroids by theintramolecular transfer of a proton from the 4β-position to the 6β-position via a dienolate intermediate. An H-bond networkin KSI consists of two catalytic residues (Tyr14 and Asp99), Tyr30, Tyr55, and a water molecule in the highly hydrophobicactive site. In this review, we summarize our current understanding of the role of the H-bond network in the activity,stability, and structure of KSI. Both mutational and structural studies suggest that the H-bond newtork could contributeto the catalysis and stability of KSI through the maintenance of the active-site geometry and the provision of structuralsupport.
Rescue of Deleterious Mutations by the Compensatory Y30F Mutation in Ketosteroid Isomerase
차형진,최관용,장도수,김연길,홍비학,우재성,김경태 한국분자세포생물학회 2013 Molecules and cells Vol.36 No.1
Proteins have evolved to compensate for detrimental mu-tations. However, compensatory mechanisms for protein defects are not well understood. Using ketosteroid isome-rase (KSI), we investigated how second-site mutations could recover defective mutant function and stability. Pre-vious results revealed that the Y30F mutation rescued the Y14F, Y55F and Y14F/Y55F mutants by increasing the catalytic activity by 23-, 3- and 1.3-fold, respectively, and the Y55F mutant by increasing the stability by 3.3 kcal/mol. To better understand these observations, we systematically investigated detailed structural and thermodynamic effects of the Y30F mutation on these mutants. Crystal structures of the Y14F/Y30F and Y14F/Y55F mutants were solved at 2.0 and 1.8 Å resolution, respectively, and compared with previoulsy solved structures of wild-type and other mutant KSIs. Structural analyses revealed that the Y30F mutation partially restored the active-site cleft of these mutant KSIs. The Y30F mutation also increased Y14F and Y14F/Y55F mutant stability by 3.2 and 4.3 kcal/mol, respectively, and the melting temperatures of the Y14F, Y55F and Y14F/Y55F mutants by 6.4°C, 5.1°C and 10.0°C, respectively. Compensatory effects of the Y30F mutation on stability might be due to improved hydrophobic interactions because removal of a hydroxyl group from Tyr30 induced local compaction by neighboring residue movement and enhanced interactions with surrounding hydrophobic residues in the active site. Taken together, our results suggest that perturbed active-site geometry recovery and favorable hydrophobic interactions mediate the role of Y30F as a second-site suppressor.
A Diagnostic Method of Control-in/out in the Glass Furnace
조진형,이세재,장도수 한국산업경영시스템학회 2006 한국산업경영시스템학회지 Vol.29 No.1
The high degree of viscosity and the non-Newtonian fluid dynamics characterizes the process inside a glass furnace. Because the temperature is fluctuating in very short time-intervals, it is hard to determine that the status of its fluctuation is stable or unstable. Usually Shewhart-chart is used to determine the control status. However because of the characteristics of the temperature fluctuations in the glass furnace it does not directly serve the purpose here. Therefore we suggest using ARIMA to diagnose control status and confirm that the method using ARIMA can be a better tool than Shewhart-chart.
용해로 운전에서 Setting고정(4M 표준화)의 효율적 방안
조진형(Jin-Hyung Cho),장도수(Do-Soo Jang),이세재(Sae-Jae Lee),서정열(Jung-Yul Suh),김이남(I-Nam Kim),정종원(Chong-Won Chung) 한국산업경영시스템학회 2007 한국산업경영시스템학회지 Vol.30 No.1
Glass melting process is influenced by both control and observation factors, where control factors include quantity and mixing ratio of raw material, the amount of fuel and air in-take. Further observation factors include temperature and pressure at each step of process inside glass melting furnace.<BR> Ambient Control is an effective means to eliminate complications from excessive variation among raw materials, or external disturbance from wide fluctuation of environment around equipments. Ambient Control uses both control and observation factors mentioned above.<BR> This study suggests an effective Proactive Control System that can enable genuine 4M standardization in glass melting furnace by applying Ambient Control.
Using Ambient Control to Prevent External Disturbances in Large-scale Furnace
조진형(Jin-Hyung Cho),장성호(Sung-Ho Chang),이세재(Sae-Jae Lee),장도수(Do-Soo Jang),서정열(Jung-Yul Suh),오현승(Hyun-Seung Oh) 한국산업경영시스템학회 2006 한국산업경영시스템학회지 Vol.29 No.2
Large glass furnaces to produce glass for CRT are housed in huge chambers. It is costly to maintain such a chamber in constant temperature, humidity, and (air) pressure. In this study, first, we show that the process of such a huge furnace, which requires the steady maintenance of high temperature, is badly affected by the ambient temperature of surrounding air. Second, an alternative process which not only maintains the relatively constant temperature dispersion around the furnace, but is also economical will be proposed. We calculate the necessary volume of air inflow in the appendix.