Studies on the roles of cavity and hydration on the enzymatic function

2014 Fiscal Year Final Research Report

• Project/Area Number: 24570186
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Biophysics
• Research Institution: Hiroshima University
• Principal Investigator: OHMAE Eiji 広島大学, 理学(系)研究科(研究院), 助教 (30284152)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Keywords: 水和 / 高圧力 / 酵素

Outline of Final Research Achievements

High-pressure stopped-flow absorbance / fluorescence spectrometer was set upped to our laboratory. The absorbance and fluorescence spectra of fluorescein were sufficiently measured but the fluorescence spectrum of intrinsic tryptophan residues of a protein was not detected by this apparatus. We measured pressure dependences of the enzymatic kinetics parameters for the wild-type and D27E mutant dihydrofolate reductases from Escherichia coli (ecDHFR) and that from deep-sea bacterium Moritella profunda (mpDHFR). The obtained Km values for ecDHFR D27E mutant and mpDHFR were less pressure-dependent than that of the wild-type ecDHFR, suggesting that the hydration around active site is important for the adaptation of DHFR to deep-sea. The results of this study clarify the molecular adaptation mechanisms of proteins from deep-sea organisms to the high-pressure environment, and give a novel guideline for the alteration of enzymes to their utilization purposes such as high-pressure condition.

Free Research Field

生物学