Investigation by atomic force microscopy of the structure of Ty3 retrotransposon particles.
Ty3, a member of the Metaviridiae family of long-terminal-repeat retrotransposons found in Saccharomyces cerevisiae, encodes homologs of retroviral Gag and Gag-Pol proteins, which, together with genomic RNA, assemble into virus-like particles (VLPs) that undergo processing and reverse transcription. The Ty3 structural proteins, capsid and nucleocapsid, contain major homology and nucleocapsid motifs similar to retrovirus capsid and nucleocapsid proteins, but Ty3 lacks a matrix-like structural domain amino terminal to capsid. Mass spectrometry analysis of Ty3 Gag3 processing products defined an acetylated Ser residue as the amino terminus of Gag3/p34, p27, and CA/p24 species and supported a model where p34 and p27 occur in phosphorylated forms. Using atomic force microscopy, VLPs were imaged from cells producing wild-type and protease and reverse transcriptase mutant Ty3. Wild-type VLPs were found to have a broad range of diameters, but the majority, if not all of the particles, exhibited arrangements of capsomeres on their surfaces which were consistent with icosahedral symmetry. Wild-type particles were in the range of 25 to 52 nm in diameter, with particles in the 42- to 52-nm diameter range consistent with T=7 symmetry. Both classes of mutant VLPs fell into a narrower range of 44 to 53 nm in diameter and appeared to be consistent with T=7 icosahedral symmetry. The smaller particles in the wild-type population likely correspond to VLPs that have progressed to reverse transcription or later stages, which do not occur in the protease and reverse transcriptase mutants. Ty3 VLPs did not undergo major external rearrangements during proteolytic maturation.
Kuznetsov
Yurii G
YG
Department of Molecular Biology, University of California, Irvine, California 92697-1700, USA.
Zhang
Min
M
Menees
Thomas M
TM
McPherson
Alexander
A
Sandmeyer
Suzanne
S
D013058:Mass Spectrometry; D018625:Microscopy, Atomic Force / Q000379:methods; D010449:Peptide Mapping; D010957:Plasmids; D012194:RNA-Directed DNA Polymerase / Q000235:genetics / Q000648:ultrastructure*; D011994:Recombinant Proteins / Q000648:ultrastructure; D018626:Retroelements; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000648:ultrastructure*
D011994:Recombinant Proteins; D018626:Retroelements; D029701:Saccharomyces cerevisiae Proteins; D012194:RNA-Directed DNA Polymerase; C496084:reverse transcriptase Ty3, S cerevisiae