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Crystal structure of a eukaryotic group II intron lariat.

Robart AR, Chan RT, Peters JK, Rajashankar KR, Toor N - Nature (2014)

Bottom Line: On the basis of structural and biochemical data, we propose that π-π' is a dynamic interaction that mediates the transition between the two steps of splicing, with η-η' serving an ancillary role.The structure also reveals a four-magnesium-ion cluster involved in both catalysis and positioning of the 5' end.Given the evolutionary relationship between group II and nuclear introns, it is likely that this active site configuration exists in the spliceosome as well.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA.

ABSTRACT
The formation of branched lariat RNA is an evolutionarily conserved feature of splicing reactions for both group II and spliceosomal introns. The lariat is important for the fidelity of 5' splice-site selection and consists of a 2'-5' phosphodiester bond between a bulged adenosine and the 5' end of the intron. To gain insight into this ubiquitous intramolecular linkage, we determined the crystal structure of a eukaryotic group IIB intron in the lariat form at 3.7 Å. This revealed that two tandem tetraloop-receptor interactions, η-η' and π-π', place domain VI in the core to position the lariat bond in the post-catalytic state. On the basis of structural and biochemical data, we propose that π-π' is a dynamic interaction that mediates the transition between the two steps of splicing, with η-η' serving an ancillary role. The structure also reveals a four-magnesium-ion cluster involved in both catalysis and positioning of the 5' end. Given the evolutionary relationship between group II and nuclear introns, it is likely that this active site configuration exists in the spliceosome as well.

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Secondary structure of P.li.LSUI2 intron crystallization construct. Tertiary interactions are indicated with Greek letters and domains are labeled with Roman numerals. Colouring of the individual domains is consistent with the overall view of the tertiary structure shown in Fig. 1.
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Figure 6: Secondary structure of P.li.LSUI2 intron crystallization construct. Tertiary interactions are indicated with Greek letters and domains are labeled with Roman numerals. Colouring of the individual domains is consistent with the overall view of the tertiary structure shown in Fig. 1.

Mentions: Group II introns are catalytic RNAs with six structural domains (Extended Data Fig. 1) that splice via two transesterification reactions. In the first step of splicing, the 2′-OH of a bulged adenosine residue is the nucleophile that attacks the 5′ splice site to generate lariat RNA4,5. In the second step, the 3′-OH of the 5′ exon attacks the 3′ splice site to form ligated exons and excised intron lariat. The highly conserved domain V (DV) forms the group II intron active site by binding catalytic metal ions9, and domain VI (DVI) contains the bulged adenosine used as the nucleophile in the first step of splicing10.


Crystal structure of a eukaryotic group II intron lariat.

Robart AR, Chan RT, Peters JK, Rajashankar KR, Toor N - Nature (2014)

Secondary structure of P.li.LSUI2 intron crystallization construct. Tertiary interactions are indicated with Greek letters and domains are labeled with Roman numerals. Colouring of the individual domains is consistent with the overall view of the tertiary structure shown in Fig. 1.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4197185&req=5

Figure 6: Secondary structure of P.li.LSUI2 intron crystallization construct. Tertiary interactions are indicated with Greek letters and domains are labeled with Roman numerals. Colouring of the individual domains is consistent with the overall view of the tertiary structure shown in Fig. 1.
Mentions: Group II introns are catalytic RNAs with six structural domains (Extended Data Fig. 1) that splice via two transesterification reactions. In the first step of splicing, the 2′-OH of a bulged adenosine residue is the nucleophile that attacks the 5′ splice site to generate lariat RNA4,5. In the second step, the 3′-OH of the 5′ exon attacks the 3′ splice site to form ligated exons and excised intron lariat. The highly conserved domain V (DV) forms the group II intron active site by binding catalytic metal ions9, and domain VI (DVI) contains the bulged adenosine used as the nucleophile in the first step of splicing10.

Bottom Line: On the basis of structural and biochemical data, we propose that π-π' is a dynamic interaction that mediates the transition between the two steps of splicing, with η-η' serving an ancillary role.The structure also reveals a four-magnesium-ion cluster involved in both catalysis and positioning of the 5' end.Given the evolutionary relationship between group II and nuclear introns, it is likely that this active site configuration exists in the spliceosome as well.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, California 92093, USA.

ABSTRACT
The formation of branched lariat RNA is an evolutionarily conserved feature of splicing reactions for both group II and spliceosomal introns. The lariat is important for the fidelity of 5' splice-site selection and consists of a 2'-5' phosphodiester bond between a bulged adenosine and the 5' end of the intron. To gain insight into this ubiquitous intramolecular linkage, we determined the crystal structure of a eukaryotic group IIB intron in the lariat form at 3.7 Å. This revealed that two tandem tetraloop-receptor interactions, η-η' and π-π', place domain VI in the core to position the lariat bond in the post-catalytic state. On the basis of structural and biochemical data, we propose that π-π' is a dynamic interaction that mediates the transition between the two steps of splicing, with η-η' serving an ancillary role. The structure also reveals a four-magnesium-ion cluster involved in both catalysis and positioning of the 5' end. Given the evolutionary relationship between group II and nuclear introns, it is likely that this active site configuration exists in the spliceosome as well.

Show MeSH