Allosteric activation of RhlB by RNase E induces partial duplex opening in substrate RNA

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• E. coli DEAD-Box helicase RhlB unwinds structured mRNA for RNA degradation by the degradosome.
• RNase E interaction allosterically stimulates RhlB's ATPase and RNA unwinding activity.
• Structural changes in RhlB upon RNase E binding and their impact on reaction rates were unclear.

Purpose of the Study:

• Investigate the structural basis for RNase E activation of RhlB.
• Characterize the effect of RNase E on RhlB's RNA binding affinity and unwinding activity.
• Explore the mechanism of RhlB activation by RNase E for different RNA substrates.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy
• RNA-centered biochemical assays
• Analysis of RNA substrates with varying topologies

Main Results:

• RNase E binding enhances RhlB's affinity for specific RNA substrates, increasing ATP turnover.
• RhlB induces conformational changes and partial opening of RNA duplexes with 5'-overhangs in the presence of RNase E, even without ATP.
• This activation mechanism is unique among DEAD-Box helicases, challenging the ATP-binding prerequisite for unwinding.

Conclusions:

• RNase E binding activates RhlB through a novel mechanism involving enhanced RNA affinity and conformational changes.
• The findings reveal a unique mode of DEAD-Box helicase activation, distinct from the canonical ATP-dependent pathway.
• This study provides new insights into the regulation of RNA metabolism and degradation in E. coli.