title: Interaction of HIV-1 gag and membranes in a cell-free system.
abstract: A coupled transcription-translation (TNT) reticulocyte lysate system was used to examine posttranslational alterations in HIV-1 Gag upon addition of Jurkat T cell membranes. Incubation of the Gag precursor protein, Pr55gag, with membranes resulted in a time-dependent alteration in Gag resulting in partial resistance to trypsin treatment. Treatment of membranes and TNT extract with apyrase or pretreatment of membranes with trypsin prevented this posttranslational alteration of Gag. In contrast, this activity was not disrupted by pretreatment of membranes with Triton X-100 at 4 degrees C, under conditions which do not solubilize raft-associated proteins. Flotation studies revealed that acquisition of trypsin-resistance was accompanied by Gag binding to membranes. The myristylation signal and nucleocapsid domain were found to mediate Gag binding to membranes. The posttranslational alteration of Gag accompanying membrane interaction may represent a conformational change, oligomerization, and/or association with or envelopment by membranes. These findings provide new clues to the stepwise process of HIV-1 assembly.
authors.affiliation: Department of Medicine, Pathology, and Molecular Mirobiology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
authors.forename: Liuzhan
authors.initials: L
authors.lastname: Yang
authors.forename: Lee
authors.initials: L
authors.lastname: Ratner
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000821:virology; D015683:Gene Products, gag / Q000378:metabolism*; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D011498:Protein Precursors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D014357:Trypsin
chemical_list: D015683:Gene Products, gag; D011498:Protein Precursors; C059767:p55 gag precursor protein, Human immunodeficiency virus 1; D000255:Adenosine Triphosphate; D014357:Trypsin