BN-PAGE analysis of DJ-1 KO dopaminergic neurons
Ontology highlight
ABSTRACT:
INSTRUMENT(S):
ORGANISM(S): Homo Sapiens (human)
TISSUE(S): Brain, Cell Culture, Diploid Cell
DISEASE(S): Parkinson's Disease
SUBMITTER: 
Christian Behrends
LAB HEAD: Christian Behrends
PROVIDER: PXD069553 | Pride | 2025-12-23
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| 20240806_SRN_014.raw | Raw | |||
| 20240806_SRN_015.raw | Raw | |||
| 20240806_SRN_016.raw | Raw | |||
| 20240806_SRN_017.raw | Raw | |||
| 20240807_SRN_034.raw | Raw |
Items per page: 5 1 - 5 of 53 |
Similar Datasets
Project description:1. PTM analysis results of DJ-1 1D-gel bands under non-reducing and reducing condition 2. HDX-MS analysis of DJ-1
2023-03-10 | PXD020690 | Pride
Project description:Mutations in the DJ-1 (Park7) gene cause autosomal recessive Parkinson's disease in humans, but the function of the DJ-1 protein is poorly characterized. In an effort to understand more about the biology of DJ-1, we performed iTRAQ analyses on subcellular fractions enriched from DJ-1 knockout rat and mouse brains. We generated iTRAQ datasets for mitochondria and cytosol enriched fractions from 6-month-old rat brains, and the cytosol enriched fraction from 14-15-month old mouse brains. Our subsequent analyses of these datasets led to our discovery that the Hexokinase 1 protein was increased in the cytosol components of both DJ-1 knockout species.
2017-10-04 | PXD005215 | Pride
Project description:Most of the redox proteomics strategies are focused on the identification and relative quantification of cysteine oxidation changes without considering the variation in the total levels of the proteins. However, the regulation of protein synthesis and protein degradation are also associated with many of the regulatory mechanisms of the cells, being therefore important to consider the changes in total protein levels in the Post Translational Modifications (PTMs) focused analyses, such as cysteine redox characterization. This aspect was only address in the cysTMTRAQ method, which combines two types of isobaric tags in the same experiment leading to a considerable increase in the costs of the analysis. Therefore, a novel integrative approach combining the SWATH-MS method with differential alkylation using non-isotopically labeled alkylating reagents (oxSWATH) is presented, thus integrating the information regarding relative cysteine oxidation with the comparative analysis of the total protein levels in a cost effective highthrouput approach. The proposed method was tested using a redox regulated protein and further applied to a comparative analysis of secretomes obtained under control or oxidative stress conditions to strengthen the importance of considering the changes in protein total levels. OxSWATH allowed to determine the relative proportion of reduced and reversible oxidized oxoforms of the proteins, and by considering total protein levels, to determine the total levels of each fraction, which are then used for comparative analysis. In this project it is presented the results from the validation of the method using the recombinant protein DJ-1.
2019-02-16 | PXD006803 | Pride
Project description:Chlamydomonas wild type CC 4533 and psb28 mutant were grown in TAP medium at 25°C and ~30 µmol photons m^-2 s^-1 on a rotary shaker. Protein complexes of three wild type (wt) and three psb28 (mut) samples were separated by BN-PAGE and cut into 36 slices. Subsequent mass spectrometry enables the analysis of protein complex migration patterns of wild type and mutant complexes.
2025-05-08 | PXD023478 | Pride
Project description:Blue Native-PAGE resolves protein complexes in their native state. When coupled with immunoblotting, it can be used to identify the presence of high molecular weight complexes at high resolution for any protein, given a suitable antibody. To identify proteins in high molecular weight complexes on a large-scale and to bypass the requirement for specific antibodies, we applied a tandem mass spectrometry (MS/MS) approach to BN-PAGE-resolved chloroplasts. Fractionation of wild-type chloroplasts into 6 bands allowed identification and label-free quantification of 1000 chloroplast proteins with native molecular weight resolution. Significantly, our approach achieves a depth of identification comparable to traditional shotgun proteomic analyses of chloroplasts, indicating much of the known chloroplast proteome is amenable to our ‘mass western’ approach to BN-PAGE. The coupling of BN-PAGE to MS/MS allows for a large-scale comparison of protein complexes between samples and the identification of proteins in high molecular weight complexes. In parallel we have analyzed chloroplasts from a leaf reticulate mutant (re-6) to demonstrate the possibility for comparative analyses. Our results provide a useful resource for the chloroplast community and this strategy is anticipated to be widely adaptable to other sub-cellular compartments.
2016-12-23 | PXD003162 | Pride
Project description:Chlamydomonas wild type CC 4533 and lpa2 mutant were grown in TAP medium at 25°C and ~30 µmol photons m^-2 s^-1 on a rotary shaker. Protein complexes of three wild type (wt) and three lpa2 (mut) samples were separated by BN-PAGE and cut into 36 slices. Subsequent mass spectrometry enables the analysis of protein complex migration patterns of wild type and mutant complexes.
2021-09-28 | PXD023443 | Pride
Project description:Photosynthesis, the fundamental process using light energy to convert CO2 to organic matter, is vital for life on Earth. It relies on capturing light through light-harvesting complexes in photosystems I and II and converting light energy into chemical energy. Composition and organization of photosystem core complexes are well conserved across evolution, and are highly susceptible to photodamage. Consequently, a large diversity of photoprotective mechanisms have evolved in photoautotrophs, finely tuned for the specific light conditions. Light Harvesting Complex protein family (LHC and LHC-like families) have acquired a dual function during evolution. Members of the LHC antenna complexes of photosystems capture light energy whereas others dissipate excess energy that cannot be harnessed for photosynthesis. This process mainly occurs through non photochemical quenching (NPQ). In this work, we focus on the LHL4 protein, which is induced by UV-B and blue light photoreceptor signaling pathways in the model green microalgae Chlamydomonas reinhardtii. Our work demonstrates that alongside established NPQ effectors, LHL4 plays a key role in photoprotection, preventing singlet oxygen accumulation in photosystem II and thereby promoting cell survival upon light stress. LHL4 protective function is distinct from that of NPQ-related proteins, since it directly binds to the transient monomeric form of the core photosystem II complex, safeguarding its integrity. Our LHL4 characterization expands our understanding of the interplay between light harvesting and photoprotection mechanisms upon light stress in photosynthetic microalgae. Here we used an approach of complexome profiling to dissect where LHL4 co-migrates in a native BN-PAGE gel and the associated proteins.
2025-01-28 | PXD055508 | Pride
Project description:Podospora anserina is an established aging model with a strong mitochondrial etiology of aging. Here we performed a complexome analysis of isolated mitochondria to study age-related changes of assembled mitochondrial protein complexes. The analysis revealed prominent age-related alterations in oxidative phosphorylation (OXPHOS) and the induction of non-mitochondrial salvage pathways.
2022-02-22 | PXD029352 | Pride
Project description:Protein complexes from isolated and crosslinked human mitochondria were separated using BN-PAGE. The gel lanes were then divided into 20 slices each, with each slice subsequently digested. The resulting peptides were analyzed, and the data from DIA runs were utilized for complexome profiling of mitochondrial complexes, including AIFM1, AK2, other proteins, and OXPHOS complexes.
2025-05-06 | PXD052821 | Pride
Project description:Here we aimed to improve native gel electrophoresis-based complexome profiling (CP) method for examination of mitochondrial (mt)DNA-/RNA-protein complexes. We compared the performance of Blue Native PAGE (BNE) and high resolution Clear Native PAGE (hrCNE). We found that hrCNE much better preserves the integrity of mtDNA-/RNA-protein complexes.To further improve the detection of mtDNA-binding proteins, we treated solubilized mitochondrial samples with DNase I prior to separation on hrCNE or BNE. To characterize the detected by hrCNE mtRNA-protein complexes, we performed CP with samples treated with RNase A. To validate the utility of the method, we performed CP using mitochondria isolated from cells treated with ethidium bromide as well as from cells recovering after this treatment. Our study not only helps to validate and unveil proteins involved in mitochondrial DNA- and RNA-related processes, but also offers a convenient and systematic way for analysing these interactions that can be used equally well for the investigation of the nucleus and other DNA-/RNA-containing cell compartments.
2023-08-11 | PXD040103 | Pride