Identification of rotenone-induced modifications in α-synuclein using affinity pull-down and tandem mass spectrometry

Hamid Mirzaei, Jeremy L. Schieler, Jean Christophe Rochet, Fred Regnier

Research output: Contribution to journalArticle

42 Citations (Scopus)

Abstract

Parkinson's disease is a movement disorder that results from a loss of dopaminergic neurons in the substantia nigra. The disease is characterized by mitochondrial dysfunction, oxidative stress, and the presence of "Lewy body" inclusions enriched with aggregated forms of α-synuclein, a presynaptic protein. Although α-synuclein is modified at various sites in Lewy bodies, it is unclear how sequence-specific posttranslational modifications modulate the aggregation of the protein in oxidatively stressed neurons. To begin to address this problem, we developed an affinity pull-down/mass spectrometry method to characterize the primary structure of histidine-tagged α-synuclein isolated from catecholaminergic neurons. Using this method, we mapped posttranslational modifications of α-synuclein from untreated neurons and neurons exposed to rotenone, an inhibitor of mitochondrial complex I. Various posttranslational modifications suggestive of oxidative damage or repair were identified in a region comprising a 20-residue stretch in the C-terminal part of the protein. The results indicate that α-synuclein is subject to discrete posttranslational modifications in neurons with impaired mitochondrial function. Our affinity pull-down/mass spectrometry method is a useful tool to examine how specific modifications of α-synuclein contribute to neurologic disorders such as Parkinson's disease.

Original languageEnglish (US)
Pages (from-to)2422-2431
Number of pages10
JournalAnalytical Chemistry
Volume78
Issue number7
DOIs
StatePublished - Apr 1 2006

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Synucleins
Rotenone
Neurons
Mass spectrometry
Proteins
Oxidative stress
Histidine
Repair
Agglomeration

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Identification of rotenone-induced modifications in α-synuclein using affinity pull-down and tandem mass spectrometry. / Mirzaei, Hamid; Schieler, Jeremy L.; Rochet, Jean Christophe; Regnier, Fred.

In: Analytical Chemistry, Vol. 78, No. 7, 01.04.2006, p. 2422-2431.

Research output: Contribution to journalArticle

Mirzaei, Hamid ; Schieler, Jeremy L. ; Rochet, Jean Christophe ; Regnier, Fred. / Identification of rotenone-induced modifications in α-synuclein using affinity pull-down and tandem mass spectrometry. In: Analytical Chemistry. 2006 ; Vol. 78, No. 7. pp. 2422-2431.
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