Probing the Structural States of Human Immunodeficiency Virus Type 1 Pr55 gag by Using Monoclonal Antibodies

Gag-FP (fluorescent protein) fusion constructs are commonly used to study human immunodeficiency virus type 1 assembly, yielding diffuse signals throughout the cytoplasm along with punctate signals routinely described as virus-like particles (VLPs) representing assembled but unprocessed Gag. However, these particles cannot be accurately described as VLPs, since fluorescence microscopy cannot provide structural resolution. We demonstrate here that the inability of a monoclonal p24 antibody to bind its cognate epitope when unprocessed Gag is assembled distinguishes VLPs from unassembled, monomeric Gag. Furthermore, we show that assembled and unassembled Gag punctate signals travel along microtubules. These monoclonal antibody studies provide a new tool for examining retroviral assembly.