title: Quantitative proteomic analysis of lentiviral vectors using 2-DE.
abstract: 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.
authors.affiliation: GENETHON, 91002 Evry, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Denard
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Rundwasser
authors.forename: Nicolas
authors.initials: N
authors.lastname: Laroudie
authors.forename: Florence
authors.initials: F
authors.lastname: Gonnet
authors.forename: Luigi
authors.initials: L
authors.lastname: Naldini
authors.forename: Marina
authors.initials: M
authors.lastname: Radrizzani
authors.forename: Anne
authors.initials: A
authors.lastname: Galy
authors.forename: Otto-Wilhelm
authors.initials: OW
authors.lastname: Merten
authors.forename: Olivier
authors.initials: O
authors.lastname: Danos
authors.forename: Fedor
authors.initials: F
authors.lastname: Svinartchouk
mesh_terms: D002460:Cell Line; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005822:Genetic Vectors / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014764:Viral Proteins / Q000378:metabolism
chemical_list: D011506:Proteins; D014764:Viral Proteins