RNA



Crystal structure of a model branchpoint–U2 snRNA duplex containing bulged adenosines


J. ANDREW  BERGLUND a1c1, MICHAEL  ROSBASH a2 and STEVE C.  SCHULTZ a3
a1 Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309, USA
a2 Howard Hughes Medical Institute, Brandeis University, Waltham, Massachusetts 02254, USA
a3 Department of Math and Sciences, Diné College, Tsaile, Arizona 86556, USA

Abstract

Bulged nucleotides play a variety of important roles in RNA structure and function, frequently forming tertiary interactions and sometimes even participating in RNA catalysis. In pre-mRNA splicing, the U2 snRNA base pairs with the intron branchpoint sequence (BPS) to form a short RNA duplex that contains a bulged adenosine that ultimately serves as the nucleophile that attacks the 5′ splice site. We have determined a 2.18-[Angstrom capital A, ring] resolution crystal structure of a self-complementary RNA designed to mimic the highly conserved yeast (Saccharomyces cerevisiae) branchpoint sequence (5′-UACUAACGUAGUA with the BPS italicized and the branchsite adenosine underlined) base paired with its complementary sequence from U2 snRNA. The structure shows a nearly ideal A-form helix from which two unpaired adenosines flip out. Although the adenosine adjacent to the branchsite adenosine is the one bulged out in the structure described here, either of these adenosines can serve as the nucleophile in mammalian but not in yeast pre-mRNA splicing. In addition, the packing of the bulged RNA helices within the crystal reveals a novel RNA tertiary interaction in which three RNA helices interact through bulged adenosines in the absence of any divalent metal ions.

(Received October 25 2000)
(Revised November 8 2000)
(Accepted January 22 2001)


Key Words: pre-mRNA splicing; RNA crystallography; three-helix RNA interaction.

Correspondence:
c1 Reprint requests to: Andy Berglund, Campus Box 215, Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309, USA; e-mail: Andy.Berglund@Colorado.edu.