The hairpin ribozyme substrate binding-domain: a highly constrained D-shaped conformation

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• The hairpin ribozyme is a catalytic RNA molecule crucial for various biological processes.
• The active conformation of the hairpin ribozyme-substrate complex requires domain interaction, but its internal geometry remains poorly understood.
• Understanding the precise structural organization is key to elucidating the catalytic mechanism.

Purpose of the Study:

• To investigate the internal geometry of the substrate-binding domain within an active hairpin ribozyme-substrate complex.
• To determine the conformational changes of the substrate-binding domain upon docking.
• To provide insights into the structural basis of hairpin ribozyme catalysis.

Main Methods:

• Utilized crosslinking and structural approaches.
• Employed molecular modeling with the constraint-satisfaction program MC-SYM.
• Analyzed the conformation of the substrate-binding domain in the context of the docked complex.

Main Results:

• The substrate-binding domain adopts a bent, D-shaped conformation in the docked state, deviating from a previously assumed straight structure.
• The helices bounding the internal loop are found to be closer than anticipated.
• This bent conformation explains the catalytic activity with circularized substrates and positions G8 near the cleavage site.

Conclusions:

• The hairpin ribozyme-substrate complex exhibits a non-linear internal geometry, specifically a bent substrate-binding domain.
• This D-shaped conformation is essential for substrate organization and catalytic activity.
• The proximity of G8 to the cleavage site, dictated by this geometry, suggests its critical role in the reaction mechanism.