Characterization of a small (25-kilodalton) derivative of the Rous sarcoma virus Gag protein competent for particle release

R. A. Weldon, John Wills

Research output: Contribution to journalArticle

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Abstract

Retroviral Gag proteins have the ability to induce budding and particle release from the plasma membrane when expressed in the absence of all of the other virus-encoded components; however, the locations of the functional domains within the Gag protein that are important for this process are poorly understood. It was shown previously that the protease sequence of the Rous sarcoma virus (RSV) Gag protein can be replaced with a foreign polypeptide, iso-1-cytochrome c from a yeast, without disrupting particle assembly (R. A. Weldon, Jr., C. R. Erdie, M. G. Oliver, and J. W. Wills, J. Virol. 64:4169- 4179, 1990). An unexpected product of the chimeric gag gene is a small, Gag- related protein named p25(C). This product was of interest because of its high efficiency of packaging into particles. The goal of the experiments described here was to determine the mechanism by which p25(C) is synthesized and packaged into particles. The results demonstrate that it is not the product of proteolytic processing of the Gag-cytochrome precursor but is derived from an unusual spliced mRNA. cDNA clones of the spliced mRNA were obtained, and each expressed a product of approximately 25 kDa, designated p25(M1), which was released into the growth medium in membrane-enclosed particles that were much lighter than authentic retrovirions as measured in sucrose density gradients. DNA sequencing revealed that the clones encode the first 180 of the 701 amino acids of the RSV Gag protein and no residues from iso-1-cytochrome c. This suggested that a domain in the carboxy-terminal half of Gag is important for the packaging of Gag proteins into dense arrays within the particles. In support of this hypothesis, particles of the correct density were obtained when a small segment from the carboxy terminus of the RSV Gag protein (residues 417 to 584) was included on the end of p25.

Original languageEnglish (US)
Pages (from-to)5550-5561
Number of pages12
JournalJournal of Virology
Volume67
Issue number9
StatePublished - Jan 1 1993

Fingerprint

Rous sarcoma virus
gag Gene Products
chemical derivatives
proteins
Cytochromes c1
cytochrome c
Product Packaging
packaging
Cytochromes a1
clones
Clone Cells
Viral Structures
Messenger RNA
cytochromes
Cytochromes
DNA Sequence Analysis
polypeptides
proteinases
plasma membrane
sequence analysis

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

Cite this

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title = "Characterization of a small (25-kilodalton) derivative of the Rous sarcoma virus Gag protein competent for particle release",
abstract = "Retroviral Gag proteins have the ability to induce budding and particle release from the plasma membrane when expressed in the absence of all of the other virus-encoded components; however, the locations of the functional domains within the Gag protein that are important for this process are poorly understood. It was shown previously that the protease sequence of the Rous sarcoma virus (RSV) Gag protein can be replaced with a foreign polypeptide, iso-1-cytochrome c from a yeast, without disrupting particle assembly (R. A. Weldon, Jr., C. R. Erdie, M. G. Oliver, and J. W. Wills, J. Virol. 64:4169- 4179, 1990). An unexpected product of the chimeric gag gene is a small, Gag- related protein named p25(C). This product was of interest because of its high efficiency of packaging into particles. The goal of the experiments described here was to determine the mechanism by which p25(C) is synthesized and packaged into particles. The results demonstrate that it is not the product of proteolytic processing of the Gag-cytochrome precursor but is derived from an unusual spliced mRNA. cDNA clones of the spliced mRNA were obtained, and each expressed a product of approximately 25 kDa, designated p25(M1), which was released into the growth medium in membrane-enclosed particles that were much lighter than authentic retrovirions as measured in sucrose density gradients. DNA sequencing revealed that the clones encode the first 180 of the 701 amino acids of the RSV Gag protein and no residues from iso-1-cytochrome c. This suggested that a domain in the carboxy-terminal half of Gag is important for the packaging of Gag proteins into dense arrays within the particles. In support of this hypothesis, particles of the correct density were obtained when a small segment from the carboxy terminus of the RSV Gag protein (residues 417 to 584) was included on the end of p25.",
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Characterization of a small (25-kilodalton) derivative of the Rous sarcoma virus Gag protein competent for particle release. / Weldon, R. A.; Wills, John.

In: Journal of Virology, Vol. 67, No. 9, 01.01.1993, p. 5550-5561.

Research output: Contribution to journalArticle

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