Plasmodium falciparum palmitome
Ontology highlight
ABSTRACT:
INSTRUMENT(S): LTQ Orbitrap Velos, instrument model
ORGANISM(S): Plasmodium Falciparum
TISSUE(S): Schizont
DISEASE(S): Plasmodium Falciparum Malaria
SUBMITTER: 
Mark Collins
PROVIDER: PRD000459 | Pride | 2012-09-24
REPOSITORIES: Pride
Dataset's files
Items per page: 5 1 - 5 of 30 |
Publications
Analysis of protein palmitoylation reveals a pervasive role in Plasmodium development and pathogenesis.
Jones Matthew L ML Collins Mark O MO Goulding David D Choudhary Jyoti S JS Rayner Julian C JC
Cell host & microbe 20120801 2
Asexual stage Plasmodium falciparum replicates and undergoes a tightly regulated developmental process in human erythrocytes. One mechanism involved in the regulation of this process is posttranslational modification (PTM) of parasite proteins. Palmitoylation is a PTM in which cysteine residues undergo a reversible lipid modification, which can regulate target proteins in diverse ways. Using complementary palmitoyl protein purification approaches and quantitative mass spectrometry, we examined p ...[more]
Similar Datasets
Project description:Biological replicate 1, technical replicate 1, SCX fraction 100 mM KH2PO4
2011-05-05 | PRD000266 | Pride
Project description:Biological replicate 2, technical replicate 1, SCX fraction 100 mM KH2PO4
2011-05-05 | PRD000267 | Pride
Project description:Plasmodium falciparum schizont proteins were extracted using SDS, digested with trypsin and phosphopeptides were enriched using IMAC. Phosphopeptides were analysed using either decision tree or data dependent neutral loss triggered ETD acquisition. All raw MS data files were processed and converted into MGF file format using Proteome Discoverer 1.1 (Thermo Scientific). A precursor filter of 600-10000 Da and a non-fragment filter were applied to ETD spectra to remove un-reacted precursor peaks, charge reduced precursor peaks, neutral losses from charge reduced precursors and FT Overtones using default settings. All ion trap spectra with less than 15 fragmentation peaks were removed and a signal to noise filter of 3 was applied to all spectra. All datasets were searched using Mascot v2.2 (Matrix Science) against a combined Human (IPI, 2010) and Plasmodium falciparum (GeneDB) sequence database (79,637 sequences) using the following search parameters: trypsin with a maximum of 3 missed cleavages, 10 ppm for MS mass tolerance, 0.5 Da for MS/MS mass tolerance, with Acetyl (Protein N-term), Oxidation (M), Deamidated (NQ), Carbamidomethyl (C) and Phospho ST set as variable modifications. ETD spectra were searched using c, z and y ion series and CID data was searched using b and y ion series. All searches used Mascot’s automated decoy database searching.
2014-04-24 | PXD000070 | Pride
Project description:Deep, strand-specific, non-polyA-selected RNA sequencing of biological replicate Plasmodium falciparum blood stage time-courses
| PRJNA246463 | ENA
Project description:Comparative phosphoproteomic analysis of SILAC labelled cultured bloodstream and procyclic form Trypansoma brucei. Phosphopeptide enrichement via SCX and TiO2, acquired on Oribtrap Velos using MSA. Triplicate biological replicate for each lifecycle stage (as unmixed, and two SILAC label swap expreiments), eight fraction injected in technical triplicate. For the SILAC labelled samples, the corresponding changes in the proteome were measured using unenriched peptides sperated by SCX. Data was processed using MaxQuant version 1.3.0.5 which incorporates the Andromeda search engine. Proteins were identified by searching a protein sequence database containingT. brucei brucei 927 annotated proteins (Version 4.0, downloaded from TriTrypDB http://www.tritrypdb.org/ supplemented with the VSG221 sequence and frequently observed contaminants (porcine trypsin, bovine serum albumins and mammalian keratins) that contains a total of 10,081 protein sequences. Search parameters specified an MS tolerance of 6 ppm, an MS/MS tolerance at 0.5 Da and full trypsin specificity, allowing for up to two missed cleavages. Carbamidomethylation of cysteine was set as a fixed modification and oxidation of methionines, N-terminal protein acetylation and N-pyroglutamate were allowed as variable modifications. Phosphoproteomic analysis included phosophorylation of serine, threonine and tyrosine as additional variable modifications. Peptides were required to be at least 7 amino acids in length and a MaxQuant score >5, with false discovery rates (FDRs) of 0.01 calculated at the levels of peptides, proteins and modification sites based on the number of hits against the reversed sequence database.
2013-07-03 | PXD000049 | Pride
Project description:Transcriptional profiling of P. falciparum cultures treated with cyclohexylamine over time (18, 25 and 30 hours post invasion) A control experiment was also set up in which P. falciparum was not treated with cyclohexylamine, and samples were taken at 18, 25 and 30 h post invasion). Background Plasmodium falciparum, the causative agent of severe human malaria, has evolved to become resistant to previously successful antimalarial chemotherapies, most notably chloroquine and the antifolates. The prevalence of resistant strains has necessitated the discovery and development of new chemical entities with novel modes-of-action. Although much effort has been invested in the creation of analogues based on existing drugs and the screening of chemical and natural compound libraries, a crucial shortcoming in current Plasmodial drug discovery efforts remains the lack of an extensive set of novel, validated drug targets. A requirement of these targets (or the pathways in which they function) is that they prove essential for parasite survival. The polyamine biosynthetic pathway, responsible for the metabolism of highly abundant amines crucial for parasite growth, proliferation and differentiation, is currently under investigation as an antimalarial target. Chemotherapeutic strategies targeting this pathway have been successfully utilized for the treatment of Trypanosomes causing West African sleeping sickness. In order to further evaluate polyamine depletion as possible antimalarial intervention, the consequences of inhibiting P. falciparum spermidine synthase (PfSpdSyn) were examined on a morphological, transcriptomic, proteomic and metabolic level. Results Morphological analysis of P. falciparum 3D7 following application of the PfSpdSyn inhibitor cyclohexylamine confirmed that parasite development was completely arrested at the early trophozoite stage. This is in contrast to untreated parasites which progressed to late trophozoites at comparable time points. Global gene expression analyses confirmed a transcriptional arrest in the parasite. Several of the differentially expressed genes mapped to the polyamine biosynthetic and associated metabolic pathways. Differential expression of corresponding parasite proteins involved in polyamine biosynthesis was also observed. Most notably, uridine phosphorylase, adenosine deaminase, lysine decarboxylase (LDC) and S-adenosylmethionine synthetase were differentially expressed at the transcript and/or protein level. Several genes in associated metabolic pathways (purine metabolism and various methyltransferases) were also affected. The specific nature of the perturbation was additionally reflected by changes in polyamine metabolite levels. Conclusions This study details the malaria parasite’s response to PfSpdSyn inhibition on the transcriptomic, proteomic and metabolic levels. The results corroborate and significantly expand previous functional genomics studies relating to polyamine depletion in this parasite. Moreover, they confirm the role of transcriptional regulation in P. falciparum, particularly in this pathway. The findings promote this essential pathway as a target for antimalarial chemotherapeutic intervention strategies. Keywords: Time course experiment in response to a drug treatment Reference design. All timepoints compared with t = 18 hours (untreated) post invasion. Two biological replicates and one technical sample per treatment and per time point. A reference design was employed for array hybridisation, utilising the URR pool described previously. All solvent-control and drug-treated sampled were hybridised to Operon slides, along with the URR. For each time point and each untreated/treated sample, three microarray slides were processed, such that a total of eighteen slides were processed in the study. Two independent cDNA samples (biological replicates) were prepared for each untreated and drug-treated sample at each time point. One of the biological replicate cDNA samples were additionally hybridised to a third slide (representing the technical replicate). GenePix results (gpr) files were generated using GenePix 6.0 (Molecular Devices) software, without normalization. For clustering analyses, results files were normalized with DNMAD (Diagnosis and Normalization for MicroArray Data) using print-tip loess. The normalized values were subsequently downloaded and analyzed with the Multiexperiment Viewer (MeV) in the TM4 software suite. Hierarchical Clustering (HCL, average linkage) was performed to estimate technical and biological variation between samples and at which point cytostasis most likely occurred for comparative purposes in downstream analyses. Intensity data for individual slides were imported into LIMMA (linear models for microarray data) in the R computing environment. Pre- and post-normalization diagnostic plots were performed using MARRAY. Data from each microarray slide was normalized using print-tip loess. Data between microarrays was normalized using rquantile normalisation. Pearson correlations were computed in ExCel to estimate variation between technical and biological replicates. Spots excluded from slide correlations and normalisation were those weighted by the limma script or flagged in the genepix results file (gpr). Additionally, spots termed Alien, Empty, Null and Operon Use Only were excluded from the correlation analyses. These spots were similarly excluded for correlations between untreated and treated samples at each time point following normalisation. Results from biological and slide replicates within each of the time points were collated, and linear models were computed to contrast gene expression between time points. A two-fold change in gene expression was used as cut-off, in conjunction with correction for false discovery (false discovery rate (FDR) = 5%). Normalised data was deposited in the Gene Expression Omnibus (GEO) database, number GSE18075. Analysis of differentially expressed genes was performed in MADIBA (Micro Array Data Interface for Biological Annotation).
2010-04-19 | E-GEOD-18075 | biostudies-arrayexpress
Project description:The palmitoyl-proteome of bovine granulosa cells (GC) cells was investigated by combining acyl-biotin exchange chemistry and quantitative mass spectrometry analysis.
2022-02-15 | PXD020547 | Pride
Project description:The palmitoyl-proteome of bovine cumulus-oocyte complexes (COCs) cells was investigated by combining acyl-biotin exchange chemistry and quantitative mass spectrometry analysis.
2022-02-15 | PXD020540 | Pride
Project description:The human malaria parasite Plasmodium falciparum employs intricate post-transcriptional regulatory mechanisms in different stages of its life cycle. Despite the importance of post-transcriptional regulation, key elements of these processes, namely RNA binding proteins (RBPs), are poorly characterized. In this study, the RNA binding properties of P. falciparum proteins were characterized including two putative members of the Bruno/CELF family of RBPs (PfCELF1 and PfCELF2), dihydrofolate reductase-thymidylate synthase (PfDHFR-TS), and adenosine deaminase (PfAda).The mRNA targets of these P. falciparum proteins were investigated by ribonomics using DNA microarrays. Two-condition ribonomic experiment, RBP vs. Mock enrichment of mRNAs. Ribonomic experimental replicates: 4-7 for each RBP, 5 for Mock. One replicate per array.
2014-10-23 | E-GEOD-56329 | biostudies-arrayexpress
Project description:MudPIT analysis of Plasmodium polysomes. TCA precipitated pellet from Plasmodium falciparum polysomes (~50ug) was solubilized in TRIS-HCl pH8.5 and Urea; reduced and alkylated. The protein suspension was digested overnight using Endoproteinase Lys-C followed by a second overnight digestion at 37°C using Trypsin. Formic acid (5% final) was added to stop the reactions. Sample was loaded on split-triple-phase fused-silica micro-capillary column and placed in-line with linear Velos pro ion-trap mass spectrometer (Thermo Scientific), coupled with quaternary Agilent 1260 series HPLC. Fully automated 10-step chromatography run (for a total of 20 hours) was carried out on the sample. Each full MS scan (400-1600 m/z) was followed by ten data-dependent MS/MS scans. The number of the micro scans was set to 1 both for MS and MS/MS. The dynamic exclusion settings used were as follows: repeat count 2; repeat duration 30s; exclusion list size 500 and exclusion duration 90 sec, the minimum signal threshold was set to 500. The MS/MS dataset was searched using SEQUEST (v.27; rev. 9) against a database of 72358 sequences, consisting of 5487 P. falciparum non-redundant proteins (downloaded from PlasmoDB on 2012-07-12), 30536 H. sapiens non-redundant proteins (downloaded from NCBI on 2012-08-27), 177 usual contaminants (such as human keratins, IgGs, and proteolytic enzymes), and, to estimate false discovery rates, 36179 randomized amino acid sequences derived from each non-redundant protein entry. To account for alkylation by CAM, 57 Da were added statically to cysteine residues and to account for the oxidation of methionine residues to methionine sulfoxide (that can occur as an artifact during sample processing) 16Da were added as a differential modification to methionine residue. Peptide/spectrum matches were sorted, selected using DTASelect/CONTRAST (v 1.9). Proteins had to be detected by 1 peptide with 2 independent spectra, leading to FDRs at the protein and spectral levels of 2.89% and 0.26%, respectively.
2013-10-31 | PXD000553 | Pride