Abstract |
We tested the hypothesis that depletion of intracellular glutathione (GSH) during heat shock results in protein thiol oxidation, thereby increasing thermal sensitivity. Depletion of GSH was accomplished using a combination of diethylmaleate and buthionine sulfoximine and protein sulfhydryls were measured using two independent methods. Chinese hamster ovary (CHO) cells were solubilized in polyacrylamide gel electrophoresis (PAGE) sample buffer containing 3-(N-maleimido-propionyl) biocytin, separated by sodium dodecyl sulfate (SDS)-PAGE, electroluted onto nitrocellulose, and visualized via avidin-alkaline phosphatase staining. A second method utilized 5,5'-dithiobis(2-nitrobenzoic acid) to measure protein solubilized in SDS. The results indicate that when CHO cells are heated at 43 degrees C GSH depletion can increase thermal sensitivity but does not cause nonspecific protein thiol oxidation at this temperature or at 37 degrees C.
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Authors | M L Freeman, M J Meredith |
Journal | Radiation research
(Radiat Res)
Vol. 117
Issue 2
Pg. 326-33
(Feb 1989)
ISSN: 0033-7587 [Print] United States |
PMID | 2493669
(Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)
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Chemical References |
- Indicators and Reagents
- Maleimides
- Proteins
- Sulfhydryl Compounds
- 3-(N-maleimidopropionyl)biocytin
- Glutathione
- Lysine
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Topics |
- Adaptation, Physiological
- Animals
- Cell Line
- Electrophoresis, Polyacrylamide Gel
- Glutathione
(physiology)
- Hot Temperature
- Indicators and Reagents
- Lysine
(analogs & derivatives)
- Maleimides
- Proteins
(analysis)
- Sulfhydryl Compounds
(analysis)
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