MudPIT analysis of HIV-1 Vpr-asssociated proteins from the permissive CEM.SS human T4-lymphoblastoid cell line

ABSTRACT: CEM.SS-iH1.Vpr cells expressing a tandem HA-FLAG- epitope-tagged HIV-1 NL4-3 Vpr (hfa-H.Vpr) under control of the Lenti-X Tet-On 3G Inducible Expression System (Clontech), and control CEM.SS cells (3x105 cells/ml, 4 liters), were induced with doxycycline (0.1ug/ml) for 9 hours. Whole cell extracts were prepared and tandem affinity purifications of Vpr and its associated proteins, from 5 grams of wet cell mass, were performed as described (Hrecka K, et al. Nature 2011; 474(7353):658-661). The Vpr-protein complexes and negative controls purified from T cells were digested with endoproteinase LysC followed by trypsin and the resulting peptide mixtures were analyzed by Multidimensional Protein Identification Technology (MudPIT) on an LTQ ion trap mass spectrometer as described (Florens L, Washburn MP. Methods Mol Biol 2006;328:159-75).

INSTRUMENT(S): LTQ

ORGANISM(S): Hiv-1 Vector Pnl4-3 (ncbitaxon:151458) Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Laurence Florens 

PROVIDER: MSV000079605 | MassIVE | Thu Mar 24 11:22:00 GMT 2016

SECONDARY ACCESSION(S): PXD003861

REPOSITORIES: MassIVE