title: Membrane binding of human immunodeficiency virus type 1 matrix protein in vivo supports a conformational myristyl switch mechanism.
abstract: The interaction of the human immunodeficiency virus (HIV) Gag protein with the plasma membrane of a cell is a critical event in the assembly of HIV particles. The matrix protein region (MA) of HIV type 1 (HIV-1) Pr55Gag has previously been demonstrated to confer membrane-binding properties on the precursor polyprotein. Both the myristic acid moiety and additional determinants within MA are essential for plasma membrane binding and subsequent particle formation. In this study, we demonstrated the myristylation-dependent membrane interaction of MA in an in vivo membrane-binding assay. When expressed within mammalian cells, MA was found both in association with cellular membranes and in a membrane-free form. In contrast, the intact precursor Pr55Gag molecule analyzed in an identical manner was found almost exclusively bound to membranes. Both membrane-bound and membrane-free forms of MA were myristylated and phosphorylated. Differential membrane binding was not due to the formation of multimers, as dimeric and trimeric forms of MA were also found in both membrane-bound and membrane-free fractions. To define the requirements for membrane binding of MA, we analyzed the membrane binding of a series of MA deletion mutants. Surprisingly, deletions within alpha-helical regions forming the globular head of MA led to a dramatic increase in overall membrane binding. The stability of the MA-membrane interaction was not affected by these deletions, and no deletion eliminated membrane binding of the molecule. These results establish that myristic acid is a primary determinant of the stability of the Gag protein-membrane interaction and provide support for the hypothesis that a significant proportion of HIV-1 MA molecules may adopt a conformation in which myristic acid is hidden and unavailable for membrane interaction.
authors.affiliation: Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2581, USA. paul.spearman@mcmail.vanderbilt.edu
authors.forename: P
authors.initials: P
authors.lastname: Spearman
authors.forename: R
authors.initials: R
authors.lastname: Horton
authors.forename: L
authors.initials: L
authors.lastname: Ratner
authors.forename: I
authors.initials: I
authors.lastname: Kuli-Zade
mesh_terms: D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000821:virology; D002498:Centrifugation; D002522:Chlorocebus aethiops; D003600:Cytosol; D010450:Endopeptidases / Q000378:metabolism; D015683:Gene Products, gag / Q000235:genetics / Q000378:metabolism*; D015488:HIV Antigens / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008561:Membrane Fusion; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism*; D010766:Phosphorylation; D011487:Protein Conformation; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D013601:T-Lymphocytes / Q000166:cytology; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus
chemical_list: D015683:Gene Products, gag; D015488:HIV Antigens; D009227:Myristic Acids; D011498:Protein Precursors; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; C054858:p17 protein, Human Immunodeficiency Virus Type 1; C059767:p55 gag precursor protein, Human immunodeficiency virus 1; D019814:Myristic Acid; D010450:Endopeptidases