1999 Fiscal Year Final Research Report Summary
Single molecule research of molecular interactions using scanning probe microscopy
| Project/Area Number |
10480185
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Biophysics
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| Research Institution | National Institute of Genetics |
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Principal Investigator |
TOKUNAGA Makio National Institute of Genetics, Structural Biology Center, Associate Professor, 構造遺伝学研究センター, 助教授 (00192659)
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| Project Period (FY) |
1998 – 1999
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| Keywords | single molecule technique / single molecule imaging / single molecule measurement / single molecule capture / motor protein / actin-myosin / loose coupling mechanism / molecular mechanism |
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| Research Abstract |
To resolve the individual mechanical events of force generation by actomyosin, we have developed a new instrument to capture and directly manipulate individual myosin subfragment-1 (S1) using a scanning probe. The single S1 molecules are visualized with a fluorescent label and the displacements during interaction with an actin bundle absorbed on a glass surface were determined by measuring the displacements of the probe. The displacements did not take place abruptly but, instead, developed in a stepwise fashion. The steps in the rising phase of displacements were regular (approximately 5.3 nm in size) and one to five steps were observed during single displacement events. The step size was constant, independent of [ATP] (0.1 and 1 ) and the temperature (20 and27℃). The mean dwell times between the steps were independent of [ATP] (3.2 and 4.8 ms at 0.1 and 1 μM, respectively), although the duration of overall displacements was greatly dependent (2 and 0.2 sec at 0.1 and 1 μM, respectively). The mean sliding velocities at the rising phase were 1.3 and 3.1 μm/s at 20 and 27℃, respectively, at 1 μM ATP, which are close to those at >1 μM ATP in a myosin-coated surface assay but >l00-fold greater than those at 1 μM ATP. These results show that a single myosin head moves along an actin filament with regular 5.3 nm steps and undergoes five steps to produce maximum displacement of approximately 30 nm for each ATP hydrolysis.
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Research Products
(10 results)
