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Quantitative proteomic analysis of lentiviral vectors using 2‐DE
Author(s) -
Denard Jérôme,
Rundwasser Stéphanie,
Laroudie Nicolas,
Gonnet Florence,
Naldini Luigi,
Radrizzani Marina,
Galy Anne,
Merten OttoWilhelm,
Danos Olivier,
Svinartchouk Fedor
Publication year - 2009
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200800747
Subject(s) - vesicular stomatitis virus , biology , proteome , cyclophilin , recombinant dna , virus , virology , group specific antigen , microbiology and biotechnology , capsid , cyclophilin a , gene , computational biology , biochemistry
Among the integrative gene therapy vectors developed to date, human immunodeficiency virus type 1 (HIV‐1)‐derived lentiviral vectors (LV) are distinguished by their capacity to infect both dividing and non‐dividing cells. Recombinant LV particles contain viral proteins necessary for their packaging, infectious and integrating functions. Like the parental HIV‐1 virus they are able to acquire various cellular proteins, but the number and localisation of these proteins are poorly characterised. In the present study we used 2‐DE followed by MALDI‐TOF to quantify the protein content of several types of vesicular stomatitis virus G‐pseudotyped LV including those that were extensively purified in the perspective of clinical gene therapy studies. A proteinase K treatment was used to distinguish between cellular proteins incorporated into virions (I‐proteins) and those co‐purified with vectors (C‐proteins). We found 10 C‐proteins and 18 I‐proteins associated with LV. Copy numbers for these core I‐proteins varied from 5 (AIP‐1/ALIX) to 280 (Cyclophilin A) per vector particle. Three novel I‐proteins, guanine nucleotide‐binding protein 2, L‐lactate dehydrogenase B chain and hnRNP core protein A1, were found. This study defines for the first time, the protein stoichiometry of infectious HIV‐1‐derived LV particles.