Aggregation of alpha-synuclein is thought to play a major role in the pathogenesis of Parkinson's disease (PD), which is characterized by the presence of intracytoplasmic Lewy bodies (LB) in the brain. alpha-Synuclein and its deletion mutants are largely unfolded proteins with random coil structures as revealed by CD spectra, fluorescence spectra, gel filtration chromatography, and ultracentrifugation. On the basis of its highly unfolded and flexible conformation, we have investigated the chaperone-like activity of alpha-synuclein in vitro. In our experiments, alpha-synuclein inhibited the aggregation of model substrates and protected the catalytic activity of alcohol dehydrogenase and rhodanese during heat stress. In addition, alpha-synuclein inhibited the initial aggregation of reduced/denatured lysozyme on the refolding pathway. Interestingly, deletion of the C-terminal regions led to the abolishment of chaperone activity, although largely unstructured conformations are maintained. Moreover, alpha-synuclein could inhibit the aggregation of various Escherichia coli cellular proteins during heat stress, and C-terminal deletion mutants could not provide any protection to these cellular proteins. Results with synthetic C-terminal peptides and C-terminal deletion mutants suggest that the second acidic repeat, (125)YEMPSEEGYQDYEPEA(140), is important for the chaperone activity of alpha-synuclein, and C-terminal deletion leads to the facilitated aggregation with the elimination of chaperone activity.

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Abstracta-Synuclein, a major constituent of Lewy bodies~LBs! in Parkinson's disease~PD!, has been implicated to play a critical role in synaptic events, such as neuronal plasticity during development, learning, and degeneration under pathological conditions, although the physiological function of a-synuclein has not yet been established. We here present biochemical evidence that recombinant a-synuclein has a chaperone-like function against thermal and chemical stress in vitro. In our experiments, a-synuclein protected glutathione S-transferase~GST! and aldolase from heatinduced precipitation, and a-lactalbumin and bovine serum albumin from dithiothreitol~DTT!-induced precipitation like other molecular chaperones. Moreover, preheating of a-synuclein, which is believed to reorganize the molecular surface of a-synuclein, increased the chaperone-like activity. Interestingly, in organic solvents, which promotes the formation of secondary structure, a-synuclein aggregated more easily than in its native condition, which eventually might abrogate the chaperone-like function of the protein. In addition, a-synuclein was also rapidly and significantly precipitated by heat in the presence of Zn 2ϩ in vitro, whereas it was not affected by the presence of Ca 2ϩ or Mg 2ϩ . Circular dichroism spectra confirmed that a-synuclein underwent conformational change in the presence of Zn 2ϩ . Taken together, our data suggest that a-synuclein could act as a molecular chaperone, and that the conformational change of the a-synuclein could explain the aggregation kinetics of a-synuclein, which may be related to the abolishment of the chaperonic-like activity.

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