Crystal structure of an RNA duplex r(gug(u)under-barcgcac)(2) with uridinebulges

The crystal structure of a nonamer RNA duplex with a uridine bulge in each strand, r(gugucgcac)(2), was determined at 1.4 Angstrom resolution. The str ucture was solved by multiple anomalous diffraction phasing method using a three-wavelength data set collected at the Advanced Protein Source and refi ned to a final R-work/R-free of 21.2%/23.4% with 33,271 independent reflect ions (Friedel pairs unmerged). The RNA duplex crystallized in the tetragona l space group P4(1)22 with two independent molecules in the asymmetric unit . The unit cell dimensions are a = b = 47.18 Angstrom and c = 80.04 Angstro m. The helical region of the nonamer adopts the A-form conformation. The ur idine bulges assume similar conformations, with uracils flipping out and pr otruding into the minor groove. The presence of the bulge induces very larg e twist angles (similar to +50 degrees) 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 bu lges, are in the C3'-endo conformation. The intrastrand phosphate-phosphate (P-P) distance of the phosphate groups flanking the bulges (similar to4.4 Angstrom) are significantly shorter than the average P-P distance in the du plex (6.0 A). This short distance between the two phosphate groups brings t he non-bridging oxygen atoms close to each other where a calcium ion is bou nd to each strand. The calcium ions in molecule 1 are well defined while th e calcium ions in molecule 2 are disordered. (C) 2001 Academic Press.