Dataset Information

Coordination of copper to the membrane-bound form of ?-synuclein.

ABSTRACT: Aggregation of the 140-amino acid protein ?-synuclein (?-syn) is linked to the development of Parkinson's disease (PD). ?-Syn is a copper binding protein with potential function as a regulator of metal-dependent redox activity. Epidemiological studies suggest that human exposure to excess copper increases the incidence of PD. ?-Syn exists in both solution and membrane-bound forms. Previous work evaluated the Cu(2+) uptake for ?-syn in solution and identified Met1-Asp2 and His50 as primary contributors to the coordination shell, with a dissociation constant of approximately 0.1 nM. When bound to the membrane bilayer, ?-syn takes on a predominantly helical conformation, which spatially separates His50 from the N-terminus of the protein and is therefore incompatible with the copper coordination geometry of the solution state. Here we use circular dichroism and electron paramagnetic resonance (continuous wave and pulsed) to evaluate the coordination of copper to the membrane-bound form of ?-syn. In this molecular environment, Cu(2+) binds exclusively to the N-terminus of the protein (Met1-Asp2) with no participation from His50. Copper does not alter the membrane-bound ?-syn conformation or enhance the release of the protein from the bilayer. The Cu(2+) affinity is similar to that identified for solution ?-syn, suggesting that copper coordination is retained in the membrane. Consideration of these results demonstrates that copper exerts its greatest conformational effect on the solution form of ?-syn.

SUBMITTER: Dudzik CG

PROVIDER: S-EPMC3544399 | biostudies-literature | 2013 Jan

REPOSITORIES: biostudies-literature

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Coordination of copper to the membrane-bound form of α-synuclein.

Dudzik Christopher G CG Walter Eric D ED Abrams Benjamin S BS Jurica Melissa S MS Millhauser Glenn L GL

Biochemistry 20121226 1

Aggregation of the 140-amino acid protein α-synuclein (α-syn) is linked to the development of Parkinson's disease (PD). α-Syn is a copper binding protein with potential function as a regulator of metal-dependent redox activity. Epidemiological studies suggest that human exposure to excess copper increases the incidence of PD. α-Syn exists in both solution and membrane-bound forms. Previous work evaluated the Cu(2+) uptake for α-syn in solution and identified Met1-Asp2 and His50 as primary contri ...[more]

PMID: 23252394

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Project description:Membrane-induced disorder-to-helix transition of α-synuclein, a presynaptic protein, has been implicated in a number of important neuronal functions as well as in the etiology of Parkinson's disease. In order to obtain structural insights of membrane-bound α-synuclein at the residue-specific resolution, we took advantage of the fact that the protein is devoid of tryptophan and incorporated single tryptophan at various residue positions along the sequence. These tryptophans were used as site-specific markers to characterize the structural and dynamical aspects of α-synuclein on the negatively charged small unilamellar lipid vesicles. An array of site-specific fluorescence readouts, such as the spectral-shift, quenching efficiency and anisotropy, allowed us to discern various features of the conformational rearrangements occurring at different locations of α-synuclein on the lipid membrane. In order to define the spatial localization of various regions of the protein near the membrane surface, we utilized a unique and sensitive indicator, namely, red-edge excitation shift (REES), which originates when a fluorophore is located in a highly ordered micro-environment. The extent of REES observed at different residue positions allowed us to directly identify the residues that are localized at the membrane-water interface comprising a thin (∼ 15 Å) layer of motionally restrained water molecules and enabled us to construct a dynamic hydration map of the protein. The combination of site-specific fluorescence readouts allowed us to unravel the intriguing molecular details of α-synuclein on the lipid membrane in a direct model-free fashion. Additionally, the combination of methodologies described here are capable of distinguishing subtle but important structural alterations of α-synuclein bound to different negatively charged lipids with varied head-group chemistry. We believe that the structural modulations of α-synuclein on the membrane could potentially be related to its physiological functions as well as to the onset of Parkinson's diseases.

Dataset Information

Coordination of copper to the membrane-bound form of ?-synuclein.

Publications

Coordination of copper to the membrane-bound form of α-synuclein.

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