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Published on: July 28, 2016
In vitro proteolytic cleavage of Gazdar murine sarcoma virus p65gag
Abstract:
Moloney murine leukemia virus, disrupted in concentrations of 0.1 to 0.5% Nonidet P-40, catalyzed the cleavage of p65, the gag gene polyprotein of the Gazdar strain of murine sarcoma virus, into polypeptides with sizes and antigenic determinants of murine leukemia virus-specified p30, p15, pp12, and p10. Cleavage performed in the presence of 0.15% Nonidet P-40 in water yielded polypeptides of approximately 40,000 (P40) and 25,000 (P25) Mr. In vitro cleavage performed in a buffered solution containing dithiothreitol in addition to 0.1% Nonidet P-40 allowed the efficient processing of P40 to p30 and a band migrating with p10. Immunoprecipitation with monospecific sera indicated that P40 contained p30 and p10, whereas P25 contained p15 and pp12 determinants. P40 and P25 are similar in size and antigenic properties to Pr40gag and Pr25gag observed in infected cells (Naso et al, J. Virol. 32:187-198, 1979).
Insights
Moloney murine leukemia virus processing of a murine sarcoma virus polyprotein was studied. The virus catalyzed cleavage into specific polypeptides, revealing key viral protein structures.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Moloney murine leukemia virus (MMLV) is a retrovirus.
- Murine sarcoma virus (MSV) encodes a gag gene polyprotein, p65.
- Understanding retroviral polyprotein processing is crucial for viral replication and pathogenesis.
Purpose of the Study:
- To investigate the in vitro cleavage of the MSV gag polyprotein (p65) by MMLV.
- To characterize the resulting polypeptide products and their antigenic determinants.
Main Methods:
- In vitro cleavage assays using MMLV and MSV p65 in varying Nonidet P-40 concentrations and buffered solutions.
- Analysis of polypeptide products by size (Mr) and immunoprecipitation with monospecific sera.
Main Results:
- MMLV catalyzed the cleavage of p65 into polypeptides antigenically related to MMLV structural proteins (p30, p15, pp12, p10).
- Cleavage in buffer with dithiothreitol yielded P40 (containing p30 and p10) and P25 (containing p15 and pp12).
- P40 and P25 resemble previously identified gag-related precursors in infected cells.
Conclusions:
- MMLV possesses enzymatic activity capable of processing the MSV gag polyprotein in vitro.
- The cleavage products share antigenic determinants with mature MMLV structural proteins.
- This study provides insights into retroviral polyprotein maturation and potential mechanisms of viral protein synthesis.
