Alpha Synuclein Interaction with Synaptic Vesicles under Glycation Conditions: Implications for Parkinson Disease and Type 2 Diabetes
DOI: Pending DOI
Synopsis
Parkinson disease and type 2 diabetes are epidemiologically associated, yet the molecular mechanisms linking these disorders remain poorly understood. Glycation, driven by elevated glucose levels and reactive carbonyl species such as methylglyoxal, can chemically modify both proteins and membrane lipids. Here we investigate whether glycation alters the interaction between alpha synuclein and synaptic vesicle membranes. Using an integrated experimental and computational approach, model vesicles were glycated in vitro and analyzed by mass spectrometry, revealing the formation of methylglyoxal derived lipid adducts. In parallel, molecular dynamics simulations were used to construct realistic neuronal membrane models. These results demonstrate that synaptic vesicle lipids can undergo glycation and establish validated membrane models for future studies of alpha synuclein membrane interactions under metabolic stress conditions.
Background
Parkinson disease is a progressive neurodegenerative disorder characterized by dopaminergic neuron loss and abnormal aggregation of alpha synuclein, a presynaptic protein involved in synaptic vesicle trafficking and neurotransmitter release (1,2). Epidemiological studies indicate that individuals with type 2 diabetes have an increased risk of developing Parkinson disease, suggesting that metabolic dysfunction may contribute to neurodegenerative processes (3).
One potential molecular mechanism linking these conditions is glycation, a non enzymatic modification caused by elevated glucose levels and reactive carbonyl species such as methylglyoxal. Glycation leads to the formation of advanced glycation end products that can alter the structure and function of proteins and lipids (4). Previous studies have shown that glycation can modify alpha synuclein and enhance its aggregation and neurotoxicity (5).
Alpha synuclein interacts directly with negatively charged phospholipid membranes and undergoes structural transitions upon membrane binding that are essential for its physiological function in synaptic vesicle regulation (6). Modifications to synaptic membrane lipids through glycation could therefore alter the physicochemical properties of membranes and disrupt protein membrane interactions.
In this study we investigate whether glycation modifies synaptic vesicle lipids and how these changes may influence alpha synuclein membrane interactions, providing insight into a possible molecular mechanism linking metabolic stress and synaptic dysfunction in Parkinson disease.
Methods
This study combines experimental biophysics and molecular dynamics simulations to investigate alpha synuclein interactions with synaptic membranes under glycation conditions.
Model vesicles were initially prepared using the negatively charged phospholipid DOPS as a simplified system to investigate lipid glycation. Vesicles were incubated with methylglyoxal to induce glycation reactions that mimic conditions associated with metabolic stress and elevated glucose levels. Lipid modifications were analyzed using electrospray ionization mass spectrometry to detect mass shifts corresponding to methylglyoxal derived lipid adducts.
Complementary computational studies were performed using molecular dynamics simulations to construct realistic neuronal membrane models. Membrane systems were built using lipid compositions representative of neuronal membranes, including DPPC, POPC, PAPC, PAPE, PUPE, DPSM, PUPS, DPGS, and PUPI together with cholesterol. All atom simulations were performed using the CHARMM36m force field (7), while coarse grained simulations were conducted using the Martini 3 model (8). Simulations were run for one microsecond to ensure structural equilibration.
Membrane organization and stability were evaluated using parameters such as area per lipid and lipid packing. These equilibrated membrane models provide a realistic platform for future studies of alpha synuclein interactions with native and glycated membranes.
Additional experimental work is currently underway using vesicle systems composed of phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, and cholesterol to better approximate synaptic vesicle membrane composition.
Results
Mass spectrometry analysis revealed clear evidence of lipid glycation following exposure of DOPS vesicles to methylglyoxal. Native DOPS vesicles displayed a characteristic peak at m z 786.52 corresponding to the expected ionized molecular mass of the lipid species. After incubation with methylglyoxal, a new peak was detected at m z 810.529, representing a mass increase consistent with the addition of a methylglyoxal derived modification. This mass shift is consistent with previously described glycation derived lipid adducts.
These findings demonstrate that synaptic vesicle lipids can undergo chemical modification through glycation under conditions associated with metabolic stress. Such modifications may alter the physicochemical properties of membranes including charge distribution, lipid packing, and bilayer organization.
In parallel, molecular dynamics simulations were used to construct and equilibrate neuronal membrane models representative of synaptic environments. Both all atom and coarse grained membrane systems were simulated for one microsecond. Analysis of the area per lipid revealed a consistent decrease relative to the initial membrane configuration during equilibration. This reduction is consistent with the ordering and condensing effects of cholesterol in neuronal membranes.The convergence of area per lipid values indicates that the membrane systems reached stable equilibrated states suitable for studying protein membrane interactions. Snapshot visualization of the equilibrated membrane further confirmed the structural stability and organization of the simulated bilayer systems.
Together, these results demonstrate that synaptic vesicle lipids are susceptible to glycation and establish validated neuronal membrane models that will enable mechanistic investigation of alpha synuclein interactions with native and chemically modified membranes.
Discussion
The present study provides experimental evidence that synaptic vesicle lipids can undergo glycation when exposed to reactive carbonyl species such as methylglyoxal. Such chemical modifications may alter the physicochemical properties of neuronal membranes and potentially influence the binding behavior of alpha synuclein, a protein central to synaptic vesicle regulation and dopamine release.
The integration of experimental and computational approaches allowed us to both detect lipid glycation and develop realistic neuronal membrane models suitable for studying protein membrane interactions. Changes in membrane composition and structure may influence alpha synuclein conformation, membrane affinity, and its ability to regulate synaptic vesicle dynamics.
These findings support the hypothesis that metabolic stress associated with type 2 diabetes could contribute to synaptic dysfunction through chemical modification of neuronal membranes and proteins. Future work will directly evaluate how glycation of membranes and alpha synuclein affects their interaction using spectroscopic and structural techniques.
Understanding how metabolic factors influence synaptic protein membrane interactions may provide insight into molecular mechanisms linking diabetes and Parkinson disease and could reveal new targets for therapeutic intervention.
Disclosures
This work was supported by a predoctoral research contract at the Universitat de les Illes Balears. The authors declare that there are no competing financial interests.
References
1. Alam P, Bousset L, Melki R, Otzen DE. Alpha synuclein oligomers and membrane interactions in Parkinson disease. Chem Rev. 2024.
2. Burré J, Sharma M, Sudhof TC. Cell biology and pathophysiology of alpha synuclein. Cold Spring Harb Perspect Med. 2018.
3. Santiago JA, Potashkin JA. Shared dysregulated pathways lead to Parkinson disease and diabetes mellitus. Trends Mol Med. 2013.
4. Singh VP, Bali A, Singh N, Jaggi AS. Advanced glycation end products and diabetic complications. Korean J Physiol Pharmacol. 2014.
5. Vicente Miranda H, Szego EM, Oliveira LMA, et al. Glycation potentiates alpha synuclein associated neurodegeneration. Acta Neuropathol. 2017.
6. Fusco G, Pape T, Stephens AD, et al. Structural basis of synaptic vesicle assembly promoted by alpha synuclein. Nat Commun. 2016.
7. Huang J, Rauscher S, Nawrocki G, et al. CHARMM36m: an improved force field for folded and intrinsically disordered proteins. Nat Methods. 2017.
8. Souza PCT, Alessandri R, Barnoud J, et al. Martini 3: a general purpose force field for coarse grained molecular dynamics. Nat Methods. 2021.