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Effect of spermidine on the RNA-A protein complex isolated from the RNA bacteriophage MS2
Abstract:
The polyamine spermidine has recently been reported to be a substantial component of the RNA phage particle. Its effect on the isolated RNA-A protein complex of the phage MS2 is investigated here. This complex infects intact Escherichia coli cells via F-pili, as does the whole phage. It is shown that the infectivity of the complex on intact E. coli cells was enhanced by incubation with spermidine. Optimal stimulation (20-fold) of the complex infectivity was achieved by incubation with 3 x 10(-4) M spermidine for 20 to 30 min at 37 degrees C. This gave a more compact structure to the complex, as could be seen by its faster sedimentation in sucrose gradients. Although spermidine and Mg2+ are known to partially replace one another in several systems, no enhancement of the infectivity of the complex, but only its considerably faster sedimentation in sucrose gradients, occurred after incubation with 3 x 10(-4) M Mg2+. Only if the Mg2+ concentration was raised by more than one order of magnitude could increased infectivity of the complex be observed. At concentrations of spermidine and Mg2+ that maximally stimulated the infectivity of the complex on intact E. coli cells, no increase in infectivity of phenol-extracted RNA to E. coli spheroplasts was detected. From these in vitro results, the role of the polyamine spermidine in the RNA phage particle for the infecting, RNA-A protein complex molecules in phage infection is discussed.
Insights
The polyamine spermidine significantly enhances the infectivity of the MS2 phage RNA-A protein complex in E. coli. This spermidine-induced enhancement is specific to the complex and not observed with isolated RNA, suggesting a role in phage entry.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- The RNA phage MS2 particle contains spermidine, a polyamine.
- The RNA-A protein complex of MS2 phage can infect E. coli cells via F-pili.
Purpose of the Study:
- To investigate the effect of spermidine on the infectivity of the isolated MS2 phage RNA-A protein complex.
- To understand the role of spermidine in phage infection mechanisms.
Main Methods:
- Incubation of the MS2 RNA-A protein complex with varying concentrations of spermidine and Mg2+.
- Assessing infectivity on intact E. coli cells.
- Analyzing complex structure via sucrose gradient sedimentation.
- Testing infectivity of phenol-extracted RNA on E. coli spheroplasts.
Main Results:
- Spermidine (3 x 10(-4) M) optimally increased complex infectivity 20-fold after 20-30 min at 37°C.
- Spermidine induced a more compact complex structure, evidenced by faster sedimentation.
- Mg2+ showed limited infectivity enhancement, requiring higher concentrations.
- Neither spermidine nor Mg2+ enhanced infectivity of isolated RNA on spheroplasts.
Conclusions:
- Spermidine plays a crucial role in enhancing the infectivity of the MS2 RNA phage RNA-A protein complex.
- The observed enhancement is likely related to the complex's interaction with the host cell surface, not RNA uptake itself.
- Spermidine's role in phage infection may involve stabilizing the complex for efficient entry via F-pili.