Microvesicles are a source of contaminating cellular proteins found in purified HIV-1 preparations.
Identification and quantitation of cellular proteins associated with HIV-1 particles are complicated by the presence of nonvirion-associated cellular proteins that copurify with virions. Many cellular proteins are associated with nonviral particles that bud from the surface of cells called microvesicles. Microvesicles band in sucrose gradients in a range of densities that includes the same density as retroviruses. To characterize these microvesicles, HIV-1-infected and uninfected human T-cell lines were propagated and virus and microvesicles were purified from clarified cell culture supernatants by sucrose density gradient centrifugation or centrifugation through 20% sucrose pads. Microvesicles were found to contain various proteins, including HLA DR and beta 2-M, and a substantial amount of RNA and DNA. The concentrations of HIV-1 p24CA, HLA DR and beta 2-microglobulin (beta 2-M) were determined by radioimmunoassay. The ratios of HIV-1 p24CA to HLA DR and beta 2-M were found to vary with respect to the HIV-1 isolate, host cell, and other factors. Electron microscopic analysis of microvesicles revealed that they consisted of particles of various sizes and morphologies. Although HIV-1 particles are known to contain some cellular proteins, microvesicles from HIV-1 infected H9 cells appeared to contain little or no HIV-1 gp120SU.
Bess
J W
JW
AIDS Vaccine Program, SAIC, National Cancer Institute-Frederick Cancer Research and Development Center, Maryland 21702-1201, USA.
Gorelick
R J
RJ
Bosche
W J
WJ
Henderson
L E
LE
Arthur
L O
LO
D016655:HIV Core Protein p24 / Q000032:analysis; D015699:HIV Envelope Protein gp120 / Q000032:analysis; D015497:HIV-1 / Q000302:isolation & purification*; D006684:HLA-DR Antigens / Q000032:analysis; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000821:virology; D015388:Organelles / Q000737:chemistry; D011506:Proteins / Q000032:analysis*; D013601:T-Lymphocytes / Q000737:chemistry / Q000821:virology; D014407:Tumor Cells, Cultured
D016655:HIV Core Protein p24; D015699:HIV Envelope Protein gp120; D006684:HLA-DR Antigens; D011506:Proteins