The Noncanonical Gag Domains p8 and n Are Critical for Assembly and Release of Mouse Mammary Tumor Virus▿

The mouse mammary tumor virus (MMTV) Gag contains the unique domains pp21, p3, p8, and n. We investigated the contribution of these domains to particle assembly and found that the region spanning the p8 and n domains is critical for shape determination and assembly. Deletion of pp21 and p3 reduced the number of released particles, but deletion of the n domain resulted in frequent formation of aberrant particles, while deletion of p8 severely impaired assembly. Further investigation of p8 revealed that both the basic and the proline-rich motifs within p8 contribute to MMTV assembly

The Mason-Pfizer Monkey Virus Internal Scaffold Domain Enables In Vitro Assembly of Human Immunodeficiency Virus Type 1 Gag

The Mason-Pfizer monkey virus (M-PMV) Gag protein possesses the ability to assemble into an immature capsid when synthesized in a reticulocyte lysate translation system. In contrast, the human immunodeficiency virus (HIV) Gag protein is incapable of assembly in parallel assays. To enable the assembly of HIV Gag, we have combined or inserted regions of M-PMV Gag into HIV Gag. By both biochemical and morphological criteria, several of these chimeric Gag molecules are capable of assembly into immature capsid-like structures in this in vitro system. Chimeric species containing large regions of M-PMV Gag fused to HIV Gag sequences failed to assemble, while species consisting of only the M-PMV p12 region, and its internal scaffold domain (ISD), fused to HIV Gag were capable of assembly, albeit at reduced kinetics compared to M-PMV Gag. The ability of the ISD to induce assembly of HIV Gag, which normally assembles at the plasma membrane, suggests a common requirement for a concentrating factor in retrovirus assembly. Despite the dramatic effect of the ISD on chimera assembly, the function of HIV Gag domains in that process was found to remain essential, since an assembly-defective mutant of HIV CA, M185A, abolished assembly when introduced into the chimera. This continued requirement for HIV Gag domain function in the assembly of chimeric molecules will allow this in vitro system to be used for the analysis of potential inhibitors of HIV immature particle assembly

Three Active Forms of Aspartic Proteinase from Mason–Pfizer Monkey Virus

Mason–Pfizer monkey virus (M-PMV) proteinase, released by the autocatalytic cleavage of Gag-Pro and Gag-Pro-Pol polypeptide precursors, catalyzes the processing of viral precursors to yield the structural proteins and enzymes of the virion. In retroviruses, usually only one proteolytically active form of proteinase exists. Here, we describe an unusual feature of M-PMV, the existence of three active forms of a retroviral proteinase with molecular masses of 17, 13, and 12 kDa as determined by mass spectroscopy. These forms arisein vitroby self-processing of a 26-kDa proteinase precursor. We have developed a process for isolation of each truncated product and demonstrate that all three forms display proteolytic activity. Amino acid analyses, as well as the determination of N- and C-terminal sequences, revealed that the N-termini of all three forms are identical, confirming thatin vitroautoprocessing of the 17-kDa form occurs at the C-terminus to yield the truncated forms. The 17-kDa form and the newly described 13-kDa form of proteinase were identified in virions collected from the rhesus monkey CMMT cell line chronically infected with M-PMV, confirming that multiple forms existin vivo

Analysis of Autoprocessing of Mason-Pfizer Monkey Virus Proteinase in Vitro Three Active Forms of Proteinase

Mason-Pfizer Monkey Virus (M-PMV, also called SRV-3) is a primate retrovirus that was first isolated from a spontaneous breast carcinoma of a female rhesus monkey Macaca mulatta.1,2 Infection of macaque species with M-PMV causes an AIDS-like disease.3,6 M-PMV is characterized by the self-assembly of the Gag, Gag-Pro and Gag-Pro-Pol precursors into intracytoplasmic particles (procapsids) within the infected cell cytoplasm. These morphogenetic properties of the virus are typical for D-type retrovirus.7,

MOESM1 of Myristoylation drives dimerization of matrix protein from mouse mammary tumor virus

Additional file 1: Figure S1. MS MALDI TOF/TOF analysis of myristoylated, myr(+), and nonmyristoylated, myr(−), MMTV MA. (A) The sample of myristoylated MA contained myr(+) MA (m/z = 13006). The sample of nonmyristoylated MA contained myr(−) MA (m/z = 12795) and MA with uncleaved initial methionine (m/z = 12927). (B) To approximately determine the minimal detectable amount of myr(−) MA in the sample of myr(+) MA, the sample of myr(+) MA was mixed with the sample of myr(−) in the ratio 1000:1 (2:0.002 mg/mL). This amount of myr(−) MA was detectable whereas the sample of myr(+) contained no detectable amount of myr(−) MA