Structural study of elements of Tetrahymena telomerase RNA stem-loop IV domain important for function

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Tetrahymena telomerase RNA (TER) has functional regions beyond the template.
• The stem-loop IV domain is critical for catalysis and ribonucleoprotein (RNP) assembly.
• Stem-loop IV binds the holoenzyme assembly protein p65 and telomerase reverse transcriptase (TERT).

Purpose of the Study:

• To determine the solution structure of TER stem-loop IV.
• To model the helical region containing the conserved GA bulge.
• To understand the structural basis for sequence conservation and protein interactions.

Main Methods:

• Nuclear Magnetic Resonance (NMR) spectroscopy.
• Residual dipolar couplings (RDCs) for structure determination.
• Biochemical analysis and structural modeling.

Main Results:

• The solution structure of loop IV was solved, and the helical region with the GA bulge was modeled.
• A conserved GA bulge with flanking C-G pairs induces a ~50-degree bend in the helix.
• A conserved C-U base pair is present at the top of the helical stem.

Conclusions:

• The structure explains sequence conservation in TER stem-loop IV.
• Protein p65 likely recognizes the helix bend in stem IV for RNP assembly.
• Binding of p65 to stem IV facilitates the positioning of loop IV for TERT interaction.