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Domain 5 binds near a highly conserved dinucleotide in the joiner linking domains 2 and 3 of a group II intron

Published online by Cambridge University Press:  01 February 1998

MIRCEA PODAR
Affiliation:
Department of Molecular Biology & Oncology, University of Texas, Southwestern Medical Center, Dallas, Texas 75235-9148, USA Present address: Biology Department, Woods Hole Oceanographic Institution, Woods Hole, Massachusetts 02543, USA.
JIN ZHUO
Affiliation:
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260-7700, USA
MINCHENG ZHANG
Affiliation:
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260-7700, USA Present address: Department of Structural Biology, Fairchild Center, Stanford University School of Medicine, Stanford, California 94305-5400, USA.
JAMES S. FRANZEN
Affiliation:
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260-7700, USA
PHILIP S. PERLMAN
Affiliation:
Department of Molecular Biology & Oncology, University of Texas, Southwestern Medical Center, Dallas, Texas 75235-9148, USA
CRAIG L. PEEBLES
Affiliation:
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260-7700, USA
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Abstract

Photocrosslinking has identified the joiner between domains 2 and 3 [J(23)] as folding near domain 5 (D5), a highly conserved helical substructure of group II introns required for both splicing reactions. D5 RNAs labeled with the photocrosslinker 4-thiouridine (4sU) reacted with highly conserved nucleotides G588 and A589 in J(23) of various intron acceptor transcripts. These conjugates retained some ribozyme function with the lower helix of D5 crosslinked to J(23), so they represent active complexes. One partner of the γ·γ′ tertiary interaction (A587·U887) is also in J(23); even though γ·γ′ is involved in step 2 of the splicing reaction, D5 has not previously been found to approach γ·γ′. Similar crosslinking patterns between D5 and J(23) were detected both before and after step 1 of the reaction, indicating that the lower helix of D5 is positioned similarly in both conformations of the active center. Our results suggest that the purine-rich J(23) strand is antiparallel to the D5 strand containing U32 and U33. Possibly, the interaction with J(23) helps position D5 correctly in the ribozyme active site; alternatively, J(23) itself might participate in the catalytic center.

Type
Research Article
Information
RNA , Volume 4 , Issue 2 , February 1998 , pp. 151 - 166
Copyright
© 1998 RNA Society

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