Structural features of membrane-bound glucocerebrosidase and α-synuclein probed by neutron reflectometry and fluorescence spectroscopy.

Abstract

Mutations in glucocerebrosidase (GCase), the enzyme deficient in Gaucher disease, are a common genetic risk factor for the development of Parkinson disease and related disorders, implicating the role of this lysosomal hydrolase in the disease etiology. A specific physical interaction exists between the Parkinson disease-related protein α-synuclein (α-syn) and GCase both in solution and on the lipid membrane, resulting in efficient enzyme inhibition. Here, neutron reflectometry was employed as a first direct structural characterization of GCase and α-syn·GCase complex on a sparsely-tethered lipid bilayer, revealing the orientation of the membrane-bound GCase. GCase binds to and partially inserts into the bilayer with its active site most likely lying just above the membrane-water interface. The interaction was further characterized by intrinsic Trp fluorescence, circular dichroism, and surface plasmon resonance spectroscopy. Both Trp fluorescence and neutron reflectometry results suggest a rearrangement of loops surrounding the catalytic site, where they extend into the hydrocarbon chain region of the outer leaflet. Taking advantage of contrasting neutron scattering length densities, the use of deuterated α-syn versus protiated GCase showed a large change in the membrane-bound structure of α-syn in the complex. We propose a model of α-syn·GCase on the membrane, providing structural insights into inhibition of GCase by α-syn. The interaction displaces GCase away from the membrane, possibly impeding substrate access and perturbing the active site. GCase greatly alters membrane-bound α-syn, moving helical residues away from the bilayer, which could impact the degradation of α-syn in the lysosome where these two proteins interact.

• Authors: Thai Leong Yap, Zhiping Jiang, Frank Heinrich, James M Gruschus, Candace M Pfefferkorn, Marilia Barros, Joseph E Curtis, Ellen Sidransky, Jennifer C Lee
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 290 Issue 2 Pg. 744-54 (Jan 09 2015) ISSN: 1083-351X [Electronic] United States
• PMID: 25429104 (Publication Type: Journal Article, Research Support, N.I.H., Intramural)
• Copyright: © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Chemical References

• Lipid Bilayers
• alpha-Synuclein
• Tryptophan
• Glucosylceramidase

Topics

• Gaucher Disease (genetics, metabolism, pathology)
• Glucosylceramidase (antagonists & inhibitors, metabolism, ultrastructure)
• Humans
• Lipid Bilayers (chemistry, metabolism)
• Mutation
• Neutron Diffraction
• Parkinson Disease (genetics, metabolism, pathology)
• Protein Binding
• Spectrometry, Fluorescence
• Surface Plasmon Resonance
• Tryptophan (chemistry)
• alpha-Synuclein (chemistry, metabolism, ultrastructure)