Crystal structure of a dimeric archaeal splicing endonuclease

Area of Science:

• Biochemistry
• Structural Biology
• Enzymology

Background:

• The Archaeoglobus fulgidus (AF) endonuclease is a homodimeric splicing endonuclease, potentially evolved from homotetrameric ancestors.
• Understanding its structure-function relationship is key to elucidating tRNA splicing mechanisms.

Purpose of the Study:

• Determine the crystal structure of the AF endonuclease.
• Identify RNA substrate binding regions and catalytic mechanisms.
• Investigate the functional implications of its higher-ordered structures.

Main Methods:

• X-ray crystallography at 2.8 Å resolution.
• Biochemical analysis.
• Structural comparison between N-terminal and C-terminal repeats.

Main Results:

• The AF endonuclease forms a homodimer with two homologous repeats: an active C-terminal and a degenerate N-terminal repeat.
• A conserved RNA-binding region for bulge-helix-bulge motifs was identified, suggesting cation-pi interactions coordinate active sites.
• A higher-ordered fibrous structure, stabilized by the N-terminal 60 residues, inhibits RNA binding and catalytic activity.

Conclusions:

• The N-terminal repeat represents an evolved, non-active endonuclease fold.
• Cation-pi interactions likely play a role in coordinating the enzyme's catalytic activity.
• The fibrous structure modulates enzyme activity, though its in vivo relevance requires further investigation.