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Defining functional groups, core structural features and inter-domain tertiary contacts essential for group II intron
1The Howard Hughes Medical Institute and Department of Biochemistry and Molecular Biophysics, 701 W. 168th Street, Room 616, Hammer Health Sciences Center, Columbia University, New York, NY 10032, USA.
The EMBO Journal
|December 8, 1998
Summary
Group II introns are self-splicing RNA molecules with key functional groups identified using nucleotide analog interference mapping (NAIM). These findings reveal critical interactions within the catalytic core, enhancing our understanding of ribozyme reactivity.
Area of Science:
- Molecular Biology
- RNA Biochemistry
Background:
- Group II introns are common ribozymes with poorly understood chemical and structural determinants of reactivity.
- Understanding these determinants is crucial for studying RNA catalysis and mobile genetic elements.
Purpose of the Study:
- To identify critical nucleotide functional groups and structural interactions within the catalytic core of Group II introns.
- To elucidate the chemical and structural basis of Group II intron self-splicing and ribozyme activity.
Main Methods:
- Nucleotide analog interference mapping (NAIM) was employed to identify essential nucleotide functional groups.
- Site-directed mutagenesis was used in conjunction with NAIM to uncover novel tertiary interactions.
Main Results:
- NAIM identified key functional groups (Rp phosphoryls, 2'-hydroxyls, guanosine exocyclic amines, adenosine N7 and N6) crucial for catalysis.
- Interference effects clustered in catalytic Domains 1 (D1) and 5 (D5), highlighting essential tetraloop-receptor interactions and a three-way junction in D1.
- A novel tertiary interaction (kappa-kappa') between D1 and D5 was discovered, stabilizing the catalytic core alongside the zeta-zeta' interaction.
Conclusions:
- The study precisely maps the chemical and structural requirements for Group II intron catalytic activity.
- The identified interactions, particularly kappa-kappa', are vital for anchoring D5 into the D1 scaffold, facilitating catalysis.
- This work provides a detailed molecular model for Group II intron ribozyme function.
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