1996 Fiscal Year Final Research Report Summary
DEVELOPMENT OF A PRESSURE-JUMP NMR APPARATUS
| Project/Area Number |
06554026
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 試験 |
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| Research Field |
Physical chemistry
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| Research Institution | DEPARTMENT OF CHEMISTRY,FACULTY OF SCIENCE,KOBE UNIVERSITY |
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Principal Investigator |
AKASAKA Kazuyuki DEPERTMENT OF CHEMISTRY,FACULTY OF SCIENCE,KOBE UNIVERSITY PROFESSOR, 理学部, 教授 (50025368)
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| Co-Investigator(Kenkyū-buntansha) |
YAMADA Hiroaki DEPERTMENT OF CHEMISTRY,FACULTY OF SCIENCE,KOBE UNIVERSITY PROFESSOR, 理学部, 助教授 (90030767)
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| Project Period (FY) |
1994 – 1996
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| Keywords | NMR / HIGH PRESSURE / PRESSURE JUMP / DENATURATION / STRUCTURAL CHANGES / PROTEIN STRUCTIRE |
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| Research Abstract |
We have designed and constructed a pressure-jump apparatus with a purpose of using it in pressure-jump NMR experiments. The first pressure-jump NMR experiment was performed for proton NMR spectrum of propanol. The jump was successfully attained from 1500 bar to 1 bar much less than a second, and the spectral change of propanol were followed with time at about 3s interval. The result showed that the signals shifted almost completely at least within 3s upon the pressure jump.
High pressure/high field NMR techniques have been developed and used for studying structure and denaturation of proteins in solution under high pressure. Thermodynamic stability of ribonuclease A (6.2mM,pH1.0,0.15M KCl, in D_2O) has been studied by ^1H NMR at 400 MHz in the pressure range of 1-2000 atm by using high pressure cell due to thermal expansion of phenylacetylene in the temperature range of 7.5-40ﾟC.The volume increase upon unfolding, DELTAV,was negative and temperature-dependent, decreasing from -10 ml/mol at 7.5ﾟC to-30ml/mol at 37ﾟC.We also found that DELTACp is dependent on pressure, decreasing from 1.79 kcal/mol K at 1 atm to 1.08 kcal/mol K at 2000 atm. An on-line high pressure quartz cell system was installed into an NMR spectrometer operating at 750MHz. This system was used to detect pressure-induced chemical shift changes of individual protons of several proteins, which demonstrated that structural changes take place within the folded conformer of a protein. In lysozyme, the hydrophobic cluster was the site of preferential compression. In other small proteins, most hydrogen bonds of the peptide NH groups were found to be shortened by pressure.
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