Crystal structure of an RNA duplex r(gugucgcac)2 with uridine bulges

The crystal structure of a nonamer RNA duplex with a uridine bulge in each strand, r(gugucgcac)2, was determined at 1.4 Å resolution. The structure was solved by multiple anomalous diffraction phasing method using a three-wavelength data set collected at the Advanced Protein Source and refined to a final Rwork/Rfree of 21.2 %/23.4 % with 33,271 independent reflections (Friedel pairs unmerged). The RNA duplex crystallized in the tetragonal space group P4122 with two independent molecules in the asymmetric unit. The unit cell dimensions are a = b = 47.18 Å and c = 80.04 Å. The helical region of the nonamer adopts the A-form conformation. The uridine bulges assume similar conformations, with uracils flipping out and protruding into the minor groove. The presence of the bulge induces very large twist angles (∼+50°) between the base-pairs flanking the bulges while causing profound kinks in the helix axis at the bulges. This severe twist and the large kink in turn produces a very narrow major groove at the middle of the molecule. The ribose sugars of the guanosines before the bulges adopt the C2′-endo conformation while the rest, including the bulges, are in the C3′-endo conformation. The intrastrand phosphate-phosphate (P-P) distance of the phosphate groups flanking the bulges (∼4.4 Å) are significantly shorter than the average P-P distance in the duplex (6.0 Å). This short distance between the two phosphate groups brings the non-bridging oxygen atoms close to each other where a calcium ion is bound to each strand. The calcium ions in molecule 1 are well defined while the calcium ions in molecule 2 are disordered.