Dataset Information

Spatial and temporal proteomic of HCMV-infected cells

ABSTRACT: Project for the study of spatial and temporal alterations in organelle proteins during HCMV infection. Two datasets are included, one for label-free quantification and one for TMT quantification. Samples were collected at 5 time points of infection (24, 48, 72, 96, and 120 hours) and subject to organelle fractionation. One non-infected sample was included for the label-free analysis, and 5 non-infected samples (one per time point) were included for the TMT dataset. The nucleus and cytosolic fractions were removed by differential centrifugation, leaving a crude organelle fraction. The crude organelle fraction was further fractionated by density gradient ultracentrifugation into sicx fractions.

INSTRUMENT(S): LTQ Orbitrap, LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (human) Human Cytomegalovirus (strain Ad169) (hhv-5) (human Herpesvirus 5)

TISSUE(S): Cell Culture, Fibroblast

SUBMITTER: Pierre Jean Beltran

LAB HEAD: Ileana Cristea

PROVIDER: PXD003925 | Pride | 2016-09-27

REPOSITORIES: Pride

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Publications

A Portrait of the Human Organelle Proteome In Space and Time during Cytomegalovirus Infection.

Jean Beltran Pierre M PM Mathias Rommel A RA Cristea Ileana M IM

Cell systems 20160915 4

The organelles within a eukaryotic host are manipulated by viruses to support successful virus replication and spread of infection, yet the global impact of viral infection on host organelles is poorly understood. Integrating microscopy, subcellular fractionation, mass spectrometry, and functional analyses, we conducted a cell-wide study of organelles in primary fibroblasts throughout the time course of human cytomegalovirus (HCMV) infection. We used label-free and isobaric-labeling proteomics t ...[more]

PMID: 27641956

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Project description:In the present study, we report for the first time a proteomic profile of Buthotus saulcyi, Odontobuthus doriae and Androctonus crassicauda scorpion venom with the aim of looking ahead and determining the structural and functional characteristics of these compounds for use as medical tools. Molecular weight determination of isolated proteins was performed in order to better describe the B. saulcyi, O. doriae and A. crassicauda raw toxin proteins by both polyacrylamide electrophoresis and two-dimensional electrophoresis. 2D-PAGE data from the B. saulcyi, O. doriae and A. crassicauda raw toxin showed a molecular weight between 3.6 and 205 kDa (for B. saulcyi), 6.6 to 205 kDa (for O. doriae) and 6.6-109 kDa (for A. crassicauda). Then 14, 14 and 21 fractions of crude toxins were isolated using HPLC and their protein content was estimated for B. saulcyi, O. doriae and A. crassicauda, respectively. SDS-PAGE analysis of selected fractions of crude toxin showed 9 protein bands with a molecular weight between 13 and 217 kDa for B. saulcyi, 10 protein bands with a molecular weight between 3.8 to182 kDa for O. doriae and 5 protein bands with a molecular weight between 5.99 and 41.65 kDa for A. crassicauda. In case of B. saulcyi, the fraction 7 (F7) showed more cytotoxicity than other isolated fractions. Subsequently, the amino acid sequencing of fraction 7 led us to two protein bands designated as p3 and p4 peptide. For O. doriae, the peptide fraction of F17, obtained from the crude venoms of O. doriae scorpion, was found to be more cytotoxic than the crude venoms and other isolated fractions. Furthermore, F5 demonstrated significant anti-proliferative and apoptotic activity. Therefore, we performed PAGE on fraction F5 and found 5 protein bands. The two protein bands, each from fraction F5 that marked as P1 and P2 were selected for amino acid sequencing. The three peptide fractions F17, obtained from the crude venoms of A. crassicauda scorpion, was found to be more cytotoxic than the crude venoms and other isolated fractions. Furthermore, F17 demonstrated significant anti-proliferative and apoptotic activity. This makes this fraction better candidate for searching the peptide that might be used for selective killing of cancerous cells. Therefore, we performed PAGE on fractionsF17, and found 2 protein bands. One protein band from fraction F17 that marked as P5 was selected for amino acid sequencing. Finally, these protein bands were removed and molecular mass and amino acid sequence analysis was performed using Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). In-silico studies of P1, P2, P3, P4 and P5 for protein sequence alignment showed the most similarity with Hemoglobin beta-2 chain protein, Chaperonin HSP60, Chrysophsin2, pheromone-bound protein 2 and Trypsin- like serine proteinase, respectively

Dataset Information

Spatial and temporal proteomic of HCMV-infected cells

Publications

A Portrait of the Human Organelle Proteome In Space and Time during Cytomegalovirus Infection.

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