RNA-binding properties of the matrix protein (p19gag) of avian sarcoma and leukemia viruses
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1Section of Biochemistry and Molecular and Cellular Biology, Cornell University, Ithaca, New York 14853.
Abstract:
We have reinvestigated the ability of the matrix protein (MA) (p19gag) of avian sarcoma and leukemia viruses to interact with RNA. Previous reports claimed on the one hand that MA can bind tightly and with a high degree of specificity to avian sarcoma and leukemia virus RNA in vitro and on the other that it cannot bind to RNA at all. We found that MA purified by any of several methods does bind to RNA, as measured by its ability to cause retention of radioactive RNA on nitrocellulose membranes in a filtration assay. However, this interaction is weak and lacks specificity. The interaction of MA with RNA was barely detectable by classical sedimentation analysis, and from this observation we estimate that the intrinsic MA-RNA association constant is ca. 10(3) M-1, at least 3 orders of magnitude smaller than the constant describing the interaction of the viral nucleocapsid protein (NC) (p12gag) with RNA, ca. 10(6) M-1. Separately purified phosphorylated and nonphosphorylated MA species bound RNA equally. We also found that MA can bind to DNA with an affinity similar to that for RNA. The large quantitative discrepancy between our results and earlier published reports can be traced in part to methods of data analysis.
Insights
The matrix protein (MA) of avian viruses weakly binds to RNA, contradicting previous claims of tight binding. This study clarifies MA
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Previous studies presented conflicting findings on the RNA-binding capabilities of the avian sarcoma and leukemia virus matrix protein (MA).
- Some research suggested MA binds RNA tightly and specifically, while others indicated no binding affinity.
- This ambiguity necessitates a re-evaluation of MA's interaction with RNA.
Purpose of the Study:
- To reinvestigate and clarify the in vitro interaction between the avian sarcoma and leukemia virus matrix protein (MA) and RNA.
- To quantify the binding affinity and specificity of MA-RNA interactions.
- To compare MA's RNA-binding properties with those of the viral nucleocapsid protein (NC).
Main Methods:
- Nitrocellulose membrane filtration assay to detect MA-RNA binding.
- Classical sedimentation analysis to assess the MA-RNA interaction strength.
- Quantification of MA-RNA association constants.
- Comparison of binding affinities for phosphorylated and nonphosphorylated MA species.
- Assessment of MA's binding affinity for DNA.
Main Results:
- Purified MA demonstrates a weak but detectable binding to RNA.
- The MA-RNA interaction lacks specificity.
- The intrinsic MA-RNA association constant is estimated at approximately 10(3) M-1.
- This affinity is significantly lower (at least 3 orders of magnitude) than that of the viral nucleocapsid protein (NC) for RNA (approximately 10(6) M-1).
- Both phosphorylated and nonphosphorylated MA bind RNA equally.
- MA exhibits similar binding affinity for DNA as for RNA.
Conclusions:
- The matrix protein (MA) of avian viruses exhibits a weak and non-specific affinity for RNA.
- MA's RNA-binding capability is considerably less than that of the viral nucleocapsid protein (NC).
- Discrepancies with prior research may stem from differences in data analysis methods.