Yeast Sis1 proximity labelling during heat shock
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive HF
ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)
SUBMITTER: 
Asif Ali
LAB HEAD: David Pincus
PROVIDER: PXD039068 | Pride | 2023-08-18
REPOSITORIES: Pride
Dataset's files
Items per page: 5 1 - 5 of 10 |
Similar Datasets
Project description:Actin, spectrin, and associated proteins form a membrane-associated periodic skeleton (MPS) in neurons. The molecular composition and functions of the MPS remain incompletely understood. Here, we combined co-immunoprecipitation and mass spectrometry analyses to identify candidate MPS-interacting proteins from cultured hippocampal neurons and adult mouse brains. In addition, we also used quantitative mass spectrometry analysis based on Tandem Mass Tag (TMT) isobaric labeling to determine how the expression levels of proteins are differentially regulated at the proteomic scale upon depletion of βII-spectrin, an important molecular component of the MPS. These results provide a systematic picture of the interactome of the MPS, and new insights into new functions of the MPS in neurons.
2022-05-04 | PXD030886 | Pride
Project description:Demand for camel milk (CM) is increasing worldwide, due to its high nutritious value and health benefits. In this study, whole CM powders were produced by spray drying (SD) at six inlet temperatures (190°C - 250°C) and by freeze drying (FD). Physicochemical and functional properties of CM powder proteins were investigated. Both treatments had negative effect on casein solubility, while whey proteins remained soluble and slightly increased its solubility with the extent of MR. The CM powders obtained at higher inlet temperatures demonstrated improved antioxidant activity. Secondary structure of whey proteins did not differ among the samples, while surface hydrophobicity of whey proteins was higher in all SD than in FD samples, suggesting only limited denaturation of camel whey proteins at higher inlet temperatures of drying. Thus, the effects of SD under the conditions applied in our study did not decrease camel whey protein solubility, while drying procedure itself regardless of temperature decreased solubility of camel milk caseins. This study provides useful insights for optimization of CM powder production.
2021-02-25 | PXD023290 | Pride
Project description:Cancers harbouring loss-of-function (LOF) alterations in tumour suppressor genes lack targeted therapies, thus alternative means to characterise gene function and identify vulnerabilities in these cancer cells are required. Here, we map the in silico genetic networks of KMT2D, a frequently mutated tumour suppressor gene, to demonstrate its utility in uncovering novel functional associations and vulnerabilities in cancer cells with tumour suppressor gene LOF alterations. In silico KMT2D networks revealed associated with histone modification, DNA replication, metabolism, and immune response. We identified synthetic lethal (SL) candidates encoding exising therapeutic targets. Analysing patient data from The Cancer Genome Atlas (TCGA) and the Personalized OncoGenomics Project (NCT021556210), we showed dysregulated pathways associated with SL candidates and elevated immune checkpoint response markers in KMT2DLOF cases, bringing forth evidence supporting KMT2D as a biomarker for immune checkpoint inhibitors. Our study presents a framework for identifying targetable vulnerabilities in cancers with tumour suppressor gene alterations.
2024-10-18 | PXD048272 | Pride
Project description:Background Plasma proteins can be modified by addition of sugar residues by non-enzymatic glycation as well as glycosylation. While glycation is a hallmark of hyperglycemia, glycosylation is a complex, enzyme-catalyzed post-translational modification by heterogeneous sugar chains and is present on a majority of plasma proteins. N-linked glycosylation occurs on asparagine residues occurring predominantly on a canonical N-glycosylation motif (Asn-X-Ser/Thr) although non-canonical N-glycosylation motifs Asn-X-Cys/Val have also been reported to be glycosylated less frequently. Albumin is the most abundant protein in human plasma whose glycation has been implicated in end-organ damage in advanced diabetes mellitus, and as such, has both diagnostic and therapeutic implications. However, albumin has long been regarded as a non-glycosylated protein owing to the absence of canonical motifs. Albumin contains two non-canonical N-glycosylation motifs, of which one was recently reported to be glycosylated with two possible attached glycans. Methods To investigate N-linked glycosylation of abundant serum proteins in greater detail, we passed healthy donor samples through a multiple affinity removal spin (MARS14) column followed by trypsin digestion of bound proteins and subsequent glycopeptide enrichment by size-exclusion chromatography (SEC) or mixed-mode anion-exchange (MAX), in parallel. Global analysis of intact glycopeptides was performed by LC-MS/MS to evaluate potential glycosylation at canonical as well as non-canonical sites. PNGase F treatment was carried out to confirm N-linked glycosylation at non-canonical sites Asn-X-Cys/Val. Glycopeptides on albumin at non-canonical sites were further characterized by MS3 fragmentation from immunoprecipitated albumin to confirm the attached glycans. To assess if the observed glycosylation was a general phenomenon, targeted analysis using parallel reaction monitoring (PRM) was carried out on an independent set of twenty human serum samples. Finally, to test whether albumin glycosylation is unique to human albumin or is a general phenomenon, bovine and rabbit serum albumins were subjected to MAX chromatography and LC-MS/MS analysis. Results We report that human albumin is modified by N-linked glycosylation at two sites, Asn68(-Glu-Val) and Asn123-(Glu-Cys), both of which occur in non-canonical N-glycosylation motifs. We detected N-glycopeptides at both sites bearing four complex mono- and bi-antennary N-linked glycans, with Hex5HexNAc4NeuAc2 and Hex5HexNAc4NeuAc1 being the most abundant glycans. These findings were validated by analyzing deglycosylated peptides and MS3-based fragmentation of glycopeptides. Glycosylation at both sites was confirmed by targeted MS in twenty donor samples. Finally, we report that the Asn residue on bovine and rabbit serum albumins that corresponds to the highly conserved Asn123 in human albumin is also N-glycosylated with complex sialylated glycans similar to those observed in human albumin. Conclusions Albumin is a conserved glycoprotein with multiple N-linked glycosylation sites. This glycosylation of albumin has potential functional implications and applications in medicine.
2024-06-16 | PXD047863 | Pride
Project description:Blood contains hundreds of proteins, reflecting ongoing cellular processes and immune reactions. Angiostrongylus vasorum infection is associated with a perturbed blood protein profile in dogs. However, the literature currently available lacks the necessary depth of analysis in order to resolve the observed pathologies in A. vasorum infections, including bleeding disorders. Using sera from 8 experimentally-infected dogs (i) before infection with A. vasorum, (ii) 34 days post-infection (p.i.; immature infection), and (iii) 75 days p.i. (mature patent infection), serum proteins were measured using liquid chromatography, tandem mass spectrometry (LC-MS/MS). For 2 dogs, serum was analyzed at days 104 and 230 p.i. additionally. A data-independent acquisition workflow was employed in order to generate quantitative data. Following computational analysis, we identified 139 up- and down-regulated proteins following infection (log2 ratio cutoff ≥ 1.0; q-value ≤ 0.05). Differences in serum profiles were most pronounced at day 75 p.i. compared to before infection. Among up-regulated proteins, chitinase 3, several saposin-like proteins, and heat shock proteins were found greatly increased (log2 fold-changes ≥ 5). Levels of pulmonary surfactant protein B were elevated on day 34 p.i. already, in the prepatent phase. Pathway enrichment revealed that complement (especially the lectin pathway) and coagulation cascades as significantly affected upon analysis of down-regulated proteins. Among them were mannan-binding lectin serine peptidases, ficolin, and coagulation factors. These results reflect the ongoing immune response and stress imposed to the lungs by the parasite. In addition, they bring new elements towards understanding the coagulopathies observed in some A. vasorum-infected dogs.
2021-01-12 | PXD016881 | Pride
Project description:A healthy existence relationship between mussels and environment relies on suitable PH, adequate oxygen and suitable temperature. Besides ocean acidification, ocean warming has been shown to cause mussel dead, a phenomenon responsible for extensive mussel dead around the word. How heat stress impacts mussel alive is poorly understood. We founded that. The superoxide dismutase (SOD) activity of two treatment groups were significantly higher than that of control group, and the activity of acid phosphatase (ACP) was significantly higher in two treatment groups than control group. The lactate dehydrogenase (LDH) activity of 26 ℃ treatment group was significantly higher than that of control group, and the LDH activity of 33 ℃ treatment group was significantly lower than that of control group. Cellular content of lysozyme (LZM) was significantly higher in two treatment groups than control group. In this study, we applied TMT quantification to investigate how Mytilus coruscus responds to heat stress (26 ℃, 33 ℃). We identified 1652 to 1878 differentially expressed proteins (DEPs) from these heat conditions, among which 897 were commonly regulated by all the two temperature and were significantly enriched in nutrients digestion and absorption (e. g. fat digestion and absorption). Furthermore, proteins associated with nutrients metabolism, signaling pathways (p53 signaling pathway and MAPK signaling pathway)and energy metabolism (e. g. TCA cycle) were impacted, suggesting that the ability of basic metabolism of Mytilus coruscus may be altered by heat stress.
2023-05-30 | PXD035618 | Pride
Project description:Proteomic analysis by combining PAGE separation, gel slicing and slice-by-slice LC-MS/MS has been frequently reported in recent years. Since the MS analysis would provide identities and quantities of the proteins along the whole lane, visualization by dye staining could be skipped to save time and labor and also in some reports assumed to improve MS identification and sensitivity. In this work, we examined the effect of CBB R-250 staining on the performance of the method and the results showed actually better results were obtained with CBB staining than without. A primary examination was firstly performed with gel bands of purified proteins, in which eight protein bands were excised, each from both the CBB-stained and unstained gel parts, and then analyzed by in-gel digestion and quantitative LC-MS/MS. Almost all the proteins were detected with higher sequence coverages and quantities from the stained gel bands than from the unstained. Then a proteomic sample of rat heart soluble proteins was examined for the complete workflow. The sample was firstly separated by nondenaturing PAGE and the gel was divided to two halves, with one CBB-stained and the other unstained. Laboratory-made tools were used to simultaneously cut duplicate lanes from each gel half and then slice each lane into about 39 pieces of the same size (1.1 mm 椋?1.1 mm 椋?1 mm thick). All the gel square pieces were analyzed in standardized procedures of in-gel digestion, peptide extraction and label-free quantitative LC-MS/MS. The results showed an average of 1434 proteins were detected in CBB-stained lanes, 40% higher than the 1013 in unstained lanes. When proteins detected in both conditions were compared, most of them were detected in higher quantities and in more gel squares with CBB staining. The comparison was also performed for SDS-PAGE and similarly advantageous results were obtained from the CBB-stained lanes, in both the detected numbers of proteins and peptides and the detected quantities and square numbers of individual proteins. The data also showed the proteins with lower molecular masses (e.g. < 30 kDa) were more benefited by the staining, probably because the dye binding helped to retain the proteins in gel matrix. In short, though dye staining is no longer a requisite when PAGE separation is followed by whole-gel LC-MS/MS analysis, CBB staining is still recommended for the better detection in proteomic analysis. All the raw and search files of the LC-MS/MS analysis, for (8*4=) 32 gel squares of HMW and LMW markers and (39*4+41*4=) 320 squares of rat heart soluble proteins (totally 704 files), as well as the gel patterns (2 files) and the summaries of the protein-level search results (3 files), are deposited in this project.
2019-02-16 | PXD011689 | Pride
Project description:LC-MS/MS analysis of O-GlcNAcylated ChREBP proteins, which were overexpressed in HEK293T cells and purified via affinity purification using Ni-NTA agarose
2017-05-04 | PXD004286 | Pride
Project description:The exocyst is a conserved octameric complex that tethers exocytic vesicles to the plasma membrane prior to fusion. Exocyst assembly and delivery mechanisms remain unclear, especially in mammalian cells. Here we tagged multiple endogenous exocyst subunits with sfGFP or Halo using Cas9 gene-editing, to create single and double knock-in lines of mammary epithelial cells, and interrogated exocyst dynamics by high-speed imaging and correlation spectroscopy. We discovered that mammalian exocyst is comprised of tetrameric subcomplexes that can associate independently with vesicles and plasma membrane and are in dynamic equilibrium with octamer and monomers. Membrane arrival times are similar for subunits and vesicles, but with a small delay (~80msec) between subcomplexes. Departure of SEC3 occurs prior to fusion, whereas other subunits depart just after fusion. About 9 exocyst complexes are associated per vesicle. These data reveal the mammalian exocyst as a remarkably dynamic two-part complex and provide important insights into assembly/disassembly mechanisms.
2018-12-05 | PXD011370 | Pride
Project description:Recombinant PDCoV spike glycoprotein was produced from Drosophile S2 cells for structural characterization by cryo electron microscopy and mass spectrometry. MS was used to assess the N-linked glycosylation profile of the purified material. We used a multiple protease digestion in combination with RP-LC-MS/MS, EThcD fragmentation on Orbitrap Fusion to characterize intact glycopeptides.
2017-10-24 | PXD007107 | Pride