1995 Fiscal Year Final Research Report Summary
IS THE RAPID-PHASE OF ENZYME CATALYTIC CYCLES ENTROPY-DRIVEN?
| Project/Area Number |
06680656
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Biophysics
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| Research Institution | KYUSHU INSTITUTE OF TECHNOLOGY |
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Principal Investigator |
KODAMA Takao KYUSHU INSTITUTE OF TECHNOLOGY,FACULTY OF COMPUTER SCIENCE AND ENGINEERING,PROFESSOR, 情報工学部, 教授 (30034200)
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| Co-Investigator(Kenkyū-buntansha) |
KOMATSU Hideyuki KYUSHU INSTITUTE OF TECHNOLOGY,FACULTY OF COMPUTER SCIENCE AND ENGINEERING,RESEA, 情報工学部, 助手 (90253567)
OKAMOTO Masahiro KYUSHU INSTITUTE OF TECHNOLOGY,FACULTY OF COMPUTER SCIENCE AND ENGINEERING,ASSOC, 情報工学部, 助教授 (40211122)
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| Project Period (FY) |
1994 – 1995
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| Keywords | thermocouple-array / stopped-flow calorimeter / chymotrypsin / simulation / intermediate steps / acylation / deacylation / peptide substrate |
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| Research Abstract |
Measurements were made of heat production accompanying hydrolysis of N-benzoyl-L-tyrosine ethyl ester (BYE) catalyzed by alpha-chymotrypsin (CT). The instrument used was essentially a stopped-flow device with a thermocouple-array as the reaction detector that can measure temperature changes < 1 mK with a response time < 5 ms [Kodama, T.& Kometani, K.(1990) Thermochim. Acta 163,105-110]. The dead time of the instrument was 13 ms. The heat records were obtained in 100 mM KCl containing 11% (v/v) acetonitril as cosolvent and analyzed by a curve-fitting procedure based on steady-state kinetic parameters estimated in separate experiments under comparable conditions (the equilibrium constant for BYE binding to CT,rate constants for acylation of CT with concomitant release of ethanol and deacylation of CT) [Berezin, I.V., Kazanskaya, N.F., & Klyosov, A.A.(1971) 15,121-124]. The enthalpy changes for these intermediate steps were taken as adjustable parmeters and their best fit values indicate that BYE binding to CT is strongly endothermic, followed by strongly exothermic CT acylation. The rate-limiting deacylation is endothermic, which makes the overall reaction almost thermally neutral.
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