A redox regulatory mechanism and a molecular link between oxidative and excitotoxic neurodegeneration have been postulated for high-affinity Na+-dependent glutamate transporters. In the present study, mutations were introduced at three cysteine residues in canine glutamate/aspartate transporter (GLAST) to investigate the functional significance of thiol groups in response to oxidation. Cys(-) GLAST, in which all cysteines were replaced by other amino acids, as well as other mutants with disruption of one of three cysteine residues, showed insoluble oligomer formation, which was considered to be due to spontaneous and excessive oxidation as observed in wild-type GLAST. The mutant transporters also showed plasma-membrane localization and glutamate-transport kinetics that were very similar to those of wild-type GLAST. Glutamate-transport activities in COS-7 cells transfected with wild-type and Cys(-) GLAST were inhibited to the same degree when cells were exposed to Hg2+ and were recovered by the addition of thiol-specific reductant dithiothreitol. These findings suggest that cysteine residues are not critical in functional expression of GLAST and the redox-sensing pathway via glutamate transporters.
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October 2002
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Research Article|
October 01 2002
Non-essential roles of cysteine residues in functional expression and redox regulatory pathways for canine glutamate/aspartate transporter based on mutagenic analysis
Satoshi TAMAHARA;
Satoshi TAMAHARA
∗Laboratory of Veterinary Clinical Pathobiology, Department of Veterinary Medical Sciences, Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo 113-8657, Japan,
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Mutsumi INABA;
Mutsumi INABA
1
∗Laboratory of Veterinary Clinical Pathobiology, Department of Veterinary Medical Sciences, Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo 113-8657, Japan,
1To whom correspondence should be addressed (e-mail ainazo@mail.ecc.u-tokyo.ac.jp).
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Kota SATO;
Kota SATO
†Department of Veterinary Internal Medicine, Faculty of Agriculture, Tottori University, Tottori 680-8553, Japan
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Naoaki MATSUKI;
Naoaki MATSUKI
∗Laboratory of Veterinary Clinical Pathobiology, Department of Veterinary Medical Sciences, Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo 113-8657, Japan,
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Yoshiaki HIKASA;
Yoshiaki HIKASA
†Department of Veterinary Internal Medicine, Faculty of Agriculture, Tottori University, Tottori 680-8553, Japan
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Ken-ichiro ONO
Ken-ichiro ONO
∗Laboratory of Veterinary Clinical Pathobiology, Department of Veterinary Medical Sciences, Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo 113-8657, Japan,
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Publisher: Portland Press Ltd
Received:
December 17 2001
Revision Received:
June 25 2002
Accepted:
June 27 2002
Accepted Manuscript online:
June 27 2002
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2002
2002
Biochem J (2002) 367 (1): 107–111.
Article history
Received:
December 17 2001
Revision Received:
June 25 2002
Accepted:
June 27 2002
Accepted Manuscript online:
June 27 2002
Citation
Satoshi TAMAHARA, Mutsumi INABA, Kota SATO, Naoaki MATSUKI, Yoshiaki HIKASA, Ken-ichiro ONO; Non-essential roles of cysteine residues in functional expression and redox regulatory pathways for canine glutamate/aspartate transporter based on mutagenic analysis. Biochem J 1 October 2002; 367 (1): 107–111. doi: https://doi.org/10.1042/bj20011843
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