Page 290 - 2014 Printable Abstract Book
P. 290
of the sample mass. Since these experiments were conducted under ultra-high vacuum conditions, we
hypothesis that the desorption of fragments from the sample into vacuum may be responsible for a large
part of the unexplained loss of DNA. To test this hypothesis, we have designed experiments to measure
in situ the absolute total mass loss due to LEE irradiation using a quartz crystal microbalance (QCM). This
experimental method is thus complementary to mass spectrometric (MS) measurements of electron
stimulated desorption and HPLC analysis of irradiated films. Here, we report results for the irradiation of
thin films of thymine. These films were deposited onto the microbalance from an effusive source. The
mass measurement was previously calibrated by HPLC on water recovered samples. The results show a
thermal-induced mass loss of thymine at room temperature (25°C) of about 2 ng/h to 4 ng/h attributed
to the slight volatility of thymine in vacuo at this temperature. Upon irradiation at 50 eV, the rate of mass
loss initially increased slightly and then decreased by between 5 and 15 fold. This change in the rate of
mass loss is indicative of structural changes occurring on the surface of the film, for example
polymerisation, induced by electron irradiation, that may prevent further fragment desorption. By water
cooling the microbalance, we can stop the thermal (background) mass loss and measure directly the initial
rate of the electron stimulated desorption. We are now moving from thymine to thymidine to eventually
reach more complex oligonuleotides. The data generated by these experiments will help to calibrate HPLC
analysis and MS measurements of fragment desorption from such films, and allow us to quantify the
absolute desorption yields.
(PS5-06) Gamma ray-induced conformational changes and improved chaperone activity of plant-
2
1
1
1
specific thioredoxin. Seung Sik Lee ; Hyoung-Woo Bai ; Jae-Young Cho ; Byung Yeoup Chung
1
Korea Atomic Energy Research Institute, Jeongeup, Korea, Republic of and Chonbuk National University,
2
Jeonju, Korea, Republic of
The heat-stable and plant-specific thioredoxin-like protein, AtTDX, is a multi-functional enzyme
present in Arabidopsis. This enzyme has been shown to act as a thioredoxin and as a molecular chaperone
depending upon its oligomeric status. In this study, we examined the effects of gamma-irradiation on the
structural and functional changes of AtTDX. Holdase chaperone activity of AtTDX reached the highest level
at 10 kGy of gamma-irradiation and then decreased further dose dependently. However, thioredoxin
activity decreased gradually with an increase in the irradiation dose. Foldase chaperone activity was not
affected much by gamma-irradiation. Electrophoresis and size exclusion chromatography (SEC) analysis
showed that AtTDX has a tendency to form gamma-ray induced high molecular weight (HMW) complexes
in a dose-dependent manner. The hydrophobicity of AtTDX increased with an increase in irradiation dose
till 20 kGy and then decreased further. Analysis of the secondary structures of AtTDX using far UV-circular
dichroism (UV-CD) spectra revealed that the irradiation significantly increased the exposure of β-sheets
and random coils with dramatic decrease in α-helices and turn elements in a dose dependent manner.
Our results suggest that gamma-irradiation may be a useful tool for increasing the chaperone holdase
activity without adverse changes in foldase chaperone activity of thioredoxin-like protein.
288 | P a g e
hypothesis that the desorption of fragments from the sample into vacuum may be responsible for a large
part of the unexplained loss of DNA. To test this hypothesis, we have designed experiments to measure
in situ the absolute total mass loss due to LEE irradiation using a quartz crystal microbalance (QCM). This
experimental method is thus complementary to mass spectrometric (MS) measurements of electron
stimulated desorption and HPLC analysis of irradiated films. Here, we report results for the irradiation of
thin films of thymine. These films were deposited onto the microbalance from an effusive source. The
mass measurement was previously calibrated by HPLC on water recovered samples. The results show a
thermal-induced mass loss of thymine at room temperature (25°C) of about 2 ng/h to 4 ng/h attributed
to the slight volatility of thymine in vacuo at this temperature. Upon irradiation at 50 eV, the rate of mass
loss initially increased slightly and then decreased by between 5 and 15 fold. This change in the rate of
mass loss is indicative of structural changes occurring on the surface of the film, for example
polymerisation, induced by electron irradiation, that may prevent further fragment desorption. By water
cooling the microbalance, we can stop the thermal (background) mass loss and measure directly the initial
rate of the electron stimulated desorption. We are now moving from thymine to thymidine to eventually
reach more complex oligonuleotides. The data generated by these experiments will help to calibrate HPLC
analysis and MS measurements of fragment desorption from such films, and allow us to quantify the
absolute desorption yields.
(PS5-06) Gamma ray-induced conformational changes and improved chaperone activity of plant-
2
1
1
1
specific thioredoxin. Seung Sik Lee ; Hyoung-Woo Bai ; Jae-Young Cho ; Byung Yeoup Chung
1
Korea Atomic Energy Research Institute, Jeongeup, Korea, Republic of and Chonbuk National University,
2
Jeonju, Korea, Republic of
The heat-stable and plant-specific thioredoxin-like protein, AtTDX, is a multi-functional enzyme
present in Arabidopsis. This enzyme has been shown to act as a thioredoxin and as a molecular chaperone
depending upon its oligomeric status. In this study, we examined the effects of gamma-irradiation on the
structural and functional changes of AtTDX. Holdase chaperone activity of AtTDX reached the highest level
at 10 kGy of gamma-irradiation and then decreased further dose dependently. However, thioredoxin
activity decreased gradually with an increase in the irradiation dose. Foldase chaperone activity was not
affected much by gamma-irradiation. Electrophoresis and size exclusion chromatography (SEC) analysis
showed that AtTDX has a tendency to form gamma-ray induced high molecular weight (HMW) complexes
in a dose-dependent manner. The hydrophobicity of AtTDX increased with an increase in irradiation dose
till 20 kGy and then decreased further. Analysis of the secondary structures of AtTDX using far UV-circular
dichroism (UV-CD) spectra revealed that the irradiation significantly increased the exposure of β-sheets
and random coils with dramatic decrease in α-helices and turn elements in a dose dependent manner.
Our results suggest that gamma-irradiation may be a useful tool for increasing the chaperone holdase
activity without adverse changes in foldase chaperone activity of thioredoxin-like protein.
288 | P a g e
