2006 Fiscal Year Final Research Report Summary
Conformational change in FALS-linked mutant Cu/Zn-SOD analyzed by monoclonal antibodies
| Project/Area Number |
17500242
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Nerve anatomy/Neuropathology
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| Research Institution | Hyogo College of Medicine |
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Principal Investigator |
FUJIWARA Noriko Hyogo College of Medicine, Faculty of Medicine, Assistant Professor, 医学部, 講師 (10368532)
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| Co-Investigator(Kenkyū-buntansha) |
SUZUKI Keiichiro Hyogo College of Medicine, Faculty of Medicine, Professor, 医学部, 教授 (70221322)
OOKAWARA Tomomi Hyogo College of Medicine, Faculty of Medicine, Assistant Professor, 医学部, 講師 (50330452)
EGUCHI Hironobu Hyogo College of Medicine, Faculty of Medicine, Research Associate, 医学部, 助手 (60351798)
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| Project Period (FY) |
2005 – 2006
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| Keywords | amyotrophic lateral sclerosis / superoxide dismutase / Cu / Zn-SOD / monoclonal antibody / oxidation / conformational change / 脳神経疾患 |
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| Research Abstract |
Amyotrophic lateral sclerosis (ALS) is a neurological disease characterized by selective motor neurons in the brain and spinal cord. Although more than 100 mutations have been identified in the Copper/Zinc-superoxide dismutase (Cu/Zn-SOD) in Familial amyotrophic lateral sclerosis (FALS), the mechanism responsible for FALS remains unclear. The finding of the present study shows that FALS causing-mutant Cu/Zn-SOD proteins (FALS mutant SODs), but not wild-type SOD, are barely detected by three monoclonal antibodies (mAbs) in Western blot analyses. ELISA for denatured FALS mutant SODs by DTT, SDS or heat treatment also showed a lowered immunoreactivity against the mAbs compared with wild-type SOD. Because all the epitopes of these mAbs are mapped within the Greek key loop (residues 102-115 in human Cu/Zn-SOD), these data uggest that different conformational changes occur in the loop between wild-type and FALS mutant SODs during the unfolding process. Circular dichroism measurements revealed that FALS mutant SODs are sensitive to denaturation by DTT, SDS or heat treatment, but these results partly but not completely explain the injurious properties of FALS mutant SODs. The findings reported herein suggest that, in addition to the instability of FALS mutant SODs, subtle conformational differences in the Greek key loop between wild-type and FALS mutant SODs during/after denaturation are involved in the etiology of FALS. In addition, recent studies suggest that oxidative damage of Cu/Zn-SOD itself has own pathogenicity in ALS. Through mass spectrometry and limited proteolysis, it was determined that the mass size of the molecule is 32 and 48, and that the modification site in Cu/Zn-SOD is Cys 111 in the Greek key loop. We demonstrated that the Cys111 is selectively oxidized to cysteine sulfinic acid (Cys-S0_2H) and to cysteine sulfonic acid (Cys-SO_3H) under mild oxidizing conditions.
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Research Products
(16 results)
