13. Ribosomes prepared from a bacterium were fractionated by sucrose density gradient centrifugation (panel i) to separate the 30S, SOS and 70S populations. When the ribosome prepuation was incubated individually with either elongation factor-G (EF-G), or a newly identified protein X. Or GTP, the profile remained unchanged. Likewise. no changes were seen in the profile when the ribosomal preparation was incubated with EF-G + GTP or protein X + GTP.
However, when the ribosomal preparation was incubated with protein X. EF.G and GTP together, itresulted in a change of profile which showed a decrease of the 70S peak area and increase in the peak areas for 30S and 50S (panel ii).

choose the option that defines a correct conclusion from the observations.
(1) Protein X is an anti-association factor which functions in the presence of EF-G and GTP
(2) Protein X is a dissociation factor which functions in the presence of EF-G and GTP
(3) Protein X binds GTP
(4) EF-G is known to bind GTP, hence it can be concluded that the effect of GTP is through EF-G and protein X does not bind GTP.

Background and Experimental Setup

Ribosomes isolated from bacteria were separated by sucrose density gradient centrifugation, yielding distinct peaks corresponding to the 30S and 50S ribosomal subunits and the intact 70S ribosome. The experiment tested the effects of elongation factor G (EF-G), a novel protein X, and GTP—individually and in combinations—on the ribosome profile.

• Incubation with EF-G, protein X, or GTP alone did not alter the ribosome profile.

• Combinations of EF-G + GTP or protein X + GTP also showed no change.

• However, incubation with protein X + EF-G + GTP together caused a decrease in the 70S peak and an increase in the 30S and 50S subunit peaks.

Interpretation of Results

The observed shift from 70S ribosomes to free 30S and 50S subunits indicates that the ribosomes are being dissociated into their individual subunits. This dissociation requires the presence of all three components: protein X, EF-G, and GTP.

• EF-G is a well-known GTPase involved in translocation during elongation and also implicated in ribosome recycling.

• The requirement of protein X together with EF-G and GTP suggests that protein X acts as a ribosome dissociation factor, facilitating the splitting of 70S ribosomes into subunits in a GTP-dependent manner.

• The lack of effect by protein X alone or with GTP indicates that protein X’s function depends on EF-G’s activity.

• The unchanged profile with EF-G + GTP alone suggests EF-G requires protein X to promote dissociation under these conditions.

Why Other Options Are Less Likely

• Anti-association factor (Option 1): Anti-association factors prevent subunits from joining but do not actively dissociate assembled 70S ribosomes. The data show active dissociation, not prevention of association.

• Protein X binds GTP (Option 3): No direct evidence from the data supports protein X binding GTP independently; the effect requires EF-G, a known GTPase.

• Effect of GTP through EF-G alone (Option 4): Since EF-G + GTP alone does not change the profile, the effect is not solely through EF-G; protein X is necessary.

Summary Table

Correct Answer

(2) Protein X is a dissociation factor which functions in the presence of EF-G and GTP

Conclusion

The combined requirement of protein X, EF-G, and GTP to dissociate 70S ribosomes into 30S and 50S subunits strongly supports the conclusion that protein X acts as a ribosome dissociation factor. This activity likely complements EF-G’s GTPase function, facilitating ribosome recycling during translation. Understanding such factors expands our knowledge of translation regulation and ribosome dynamics in bacteria.