Screening of Crucial Cytosolicproteins Interconnecting the Endoplasmic Reticulum and Mitochondria in Parkinson's Disease and the Impact of Anti-Parkinson Drugs in the Preservation of Organelle Connectivity

Abstract

Mitochondrial dysfunction is well-established in Parkinson's disease (PD); however, its dysfunctions associating with cell organelle connectivity remain unknown. We aimed to establish the crucial cytosolic protein involved in organelle connectivity between mitochondria and the endopalmic reticulum (ER) through a computational approach by constructing an organelle protein network to extract functional clusters presenting the crucial PD protein connecting organelles. Then, we assessed the influence of anti-parkinsonism drugs (n = 35) on the crucial protein through molecular docking and molecular dynamic simulation and further validated its gene expression in PD participants under, istradefylline (n = 25) and amantadine (n = 25) treatment. Based on our investigation, D-aspartate oxidase (DDO )protein was found to be the critical that connects both mitochondria and the ER. Further, molecular docking showed that istradefylline has a high affinity (-9.073 kcal/mol) against DDO protein, which may disrupt mitochondrial-ER connectivity. While amantadine (-4.53 kcal/mol) shows negligible effects against DDO that contribute to conformational changes in drug binding, Successively, DDO gene expression was downregulated in istradefylline-treated PD participants, which elucidated the likelihood of an istradefylline off-target mechanism. Overall, our findings illuminate the off-target effects of anti-parkinsonism medications on DDO protein, enabling the recommendation of off-target-free PD treatments.

Keywords: Parkinson’s disease; mitochondria-associated membranes (MAMs); mitochondrial-ER; neuroinflammation; systems biology.

Figure 1

Comprehensive overview of the current investigation: Section-A: collection of organelle-specific proteins of the endoplasmic reticulum (ER), mitochondria, and cytosol from various databases and conversion of the collected proteins into official symbols in order to build a protein interaction network. The top three clusters were extracted from the protein network using MCODE clustering. Simultaneously, meta-gene expression analysis was carried out with PD datasets retrieved from the NCBI-GEO datasets. The differentially expressed genes from the meta-analysis were mapped to the top three clusters to select clusters influenced by PD pathogenesis (termed PD cluster) and screened to select the cluster presenting proteins localized to the mitochondria, endoplasmic reticulum, and cytosol. Then, protein shortest path analysis was conducted to discover the crucial cytosolic protein connecting the mitochondria and endoplasmic reticulum (ER). Further, molecular docking and molecular dynamic (MD) simulation were performed to evaluate the impact of anti-Parkinson’s disease drugs on the crucial cytosolic protein. Section-B: To validate our finding, qPCR was used to measure gene expression in peripheral blood mononuclear cells (PBMC) of PD drug-treated groups and controls for the selected gene, and the significance was assessed by statistical analysis.

Figure 2

PD Cluster with mitochondrial, ER, and cytosolic proteins. A node represented in orange is the endoplasmic reticulum; a green-colored node represents mitochondrial proteins; and a blue-colored node corresponds to the proteins localized in the cytosol. The red-colored highlighted edges show the inter-cell organelle interaction via protein signal.

Figure 3

Molecular enrichment analysis of selected PD cluster.

Figure 4

Ligand interaction diagram for the docked anti-Parkinsonism drugs (A) istradefylline drug contributes high affinity with DDO cytosol protein, forming a significant number of hydrogen bonds; and (B) amantadine drug contributes less affinity with no hydrogen bond formation with the DDO protein.

Figure 5

Molecular dynamic simulation of ligand free DDO (colored: black), DDO_istradefylline (colored: red), and (3) DDO_amantadine (colored: green) complexes. (A) root-mean-square deviation (RMSD), (B) root-mean-square fluctuation (RMSF), (C) solvent accessible surface area (SASA), (D) radius of gyration (Rg), and (E) H-bonds.

Figure 6

Gene expression analysis of DDO gene with istradefylline and amantadine.The relative expression of DDO shows significantly decreased DDO gene expression when treated with istradefylline. (**) significant changes were observed in the PD participant treated with istradefylline and amantadine compared to the control.