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 A 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 R(work)/R(free) of 21.2 %/23.4 % with 33,271 independent reflections (Friedel pairs unmerged). The RNA duplex crystallized in the tetragonal space group P4(1)22 with two independent molecules in the asymmetric unit. The unit cell dimensions are a=b=47.18 A and c=80.04 A. 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 (approximately +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 bulges, are in the C3'-endo conformation. The intrastrand phosphate-phosphate (P-P) distance of the phosphate groups flanking the bulges (approximately 4.4 A) are significantly shorter than the average P-P distance in the duplex (6.0 A). 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.

[1]  N. Usman,et al.  X-ray crystallographic observation of "in-line" and "adjacent" conformations in a bulged self-cleaving RNA/DNA hybrid. , 2001, RNA.

[2]  M. Sundaralingam,et al.  Two crystal forms of helix II of Xenopus laevis 5S rRNA with a cytosine bulge. , 2000, RNA.

[3]  I. Tinoco,et al.  Solution structure and metal-ion binding of the P4 element from bacterial RNase P RNA. , 2000, RNA.

[4]  M. Sundaralingam,et al.  Crystal structure of an adenine bulge in the RNA chain of a DNA.RNA hybrid, d(CTCCTCTTC).r(gaagagagag). , 2000, Journal of molecular biology.

[5]  D. Patel,et al.  RNA bulges as architectural and recognition motifs. , 2000, Structure.

[6]  T. Steitz,et al.  The structure of the HIV-1 RRE high affinity rev binding site at 1.6 A resolution. , 2000, Journal of molecular biology.

[7]  C. Ehresmann,et al.  The crystal structure of the dimerization initiation site of genomic HIV-1 RNA reveals an extended duplex with two adenine bulges. , 1999, Structure.

[8]  J. Wedekind,et al.  Crystal structure of a lead-dependent ribozyme revealing metal binding sites relevant to catalysis , 1999, Nature Structural Biology.

[9]  Russ Miller,et al.  The design and implementation of SnB version 2.0 , 1999 .

[10]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[11]  T. Steitz,et al.  A 1.3-A resolution crystal structure of the HIV-1 trans-activation response region RNA stem reveals a metal ion-dependent bulge conformation. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[12]  T. Steitz,et al.  Metals, Motifs, and Recognition in the Crystal Structure of a 5S rRNA Domain , 1997, Cell.

[13]  J. Murray,et al.  The three-dimensional structures of two complexes between recombinant MS2 capsids and RNA operator fragments reveal sequence-specific protein-RNA interactions. , 1997, Journal of molecular biology.

[14]  E. Westhof,et al.  The RNA binding site of S8 ribosomal protein of Escherichia coli: Selex and hydroxyl radical probing studies. , 1997, RNA.

[15]  N. Pace,et al.  In vitro selection of RNase P RNA reveals optimized catalytic activity in a highly conserved structural domain. , 1996, RNA.

[16]  C. Kundrot,et al.  Crystal Structure of a Group I Ribozyme Domain: Principles of RNA Packing , 1996, Science.

[17]  N. Usman,et al.  Crystal structures of an A-form duplex with single-adenosine bulges and a conformational basis for site-specific RNA self-cleavage. , 1996, Chemistry & biology.

[18]  D. Lilley.  Kinking of DNA and RNA by base bulges. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Philip N. Borer,et al.  Proton NMR and structural features of a 24-nucleotide RNA hairpin. , 1995, Biochemistry.

[20]  Collaborative Computational,et al.  The CCP4 suite: programs for protein crystallography. , 1994, Acta crystallographica. Section D, Biological crystallography.

[21]  R D Klausner,et al.  The interaction between the iron-responsive element binding protein and its cognate RNA is highly dependent upon both RNA sequence and structure. , 1993, Nucleic acids research.

[22]  J. Karn,et al.  Recognition of the high affinity binding site in rev-response element RNA by the human immunodeficiency virus type-1 rev protein. , 1992, Nucleic acids research.

[23]  D. Patel,et al.  Comparative NMR study of A(n)-bulge loops in DNA duplexes: intrahelical stacking of A, A-A, and A-A-A bulge loops. , 1992, Biochemistry.

[24]  D A LeBlanc,et al.  Thermodynamic characterization of deoxyribooligonucleotide duplexes containing bulges. , 1991, Biochemistry.

[25]  M. Singh,et al.  HIV‐1 tat protein stimulates transcription by binding to a U‐rich bulge in the stem of the TAR RNA structure. , 1990, The EMBO journal.

[26]  K. Maskos,et al.  NMR studies of a deoxyribodecanucleotide containing an extrahelical thymidine surrounded by an oligo(dA).oligo(dT) tract. , 1990, Biochemistry.

[27]  D. Patel,et al.  Conformational transitions in thymidine bulge-containing deoxytridecanucleotide duplexes. Role of flanking sequence and temperature in modulating the equilibrium between looped out and stacked thymidine bulge states. , 1990, The Journal of biological chemistry.

[28]  C. E. Longfellow,et al.  Thermodynamic and spectroscopic study of bulge loops in oligoribonucleotides. , 1990, Biochemistry.

[29]  D. Gorenstein,et al.  Two-dimensional 1H and 31P NMR spectra and restrained molecular dynamics structure of an extrahelical adenosine tridecamer oligodeoxyribonucleotide duplex. , 1989, Biochemistry.

[30]  D. Crothers,et al.  Conformation of a bulge‐containing oligomer from a hot‐spot sequence by NMR and energy minimization , 1989, Biopolymers.

[31]  C R Woese,et al.  Evidence for several higher order structural elements in ribosomal RNA. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[32]  D. Patel,et al.  Conformational transitions in cytidine bulge-containing deoxytridecanucleotide duplexes: extra cytidine equilibrates between looped out (low temperature) and stacked (elevated temperature) conformations in solution. , 1989, Biochemistry.

[33]  R Lavery,et al.  The definition of generalized helicoidal parameters and of axis curvature for irregular nucleic acids. , 1988, Journal of biomolecular structure & dynamics.

[34]  G. A. van der Marel,et al.  Bulge-out structures in the single-stranded trimer AUA and in the duplex (CUGGUGCGG).(CCGCCCAG). A model-building and NMR study. , 1988, Nucleic acids research.

[35]  D. Crothers,et al.  Structural model for an oligonucleotide containing a bulged guanosine by NMR and energy minimization. , 1988, Biochemistry.

[36]  H. Elst,et al.  Extra thymidine stacks into the d(CTGGTGCGG).d(CCGCCCAG) duplex. An NMR and model-building study. , 1988, Nucleic acids research.

[37]  D. Patel,et al.  Extrahelical adenosine stacks into right-handed DNA: solution conformation of the d(C-G-C-A-G-A-G-C-T-C-G-C-G) duplex deduced from distance geometry analysis of nuclear Overhauser effect spectra. , 1986, Biochemistry.

[38]  Ignacio Tinoco,et al.  Unpaired cytosine in the deoxyoligonucleotide duplex dCA3CA3G.cntdot.dCT6G is outside of the helix , 1983 .

[39]  D. Patel,et al.  Extra adenosine stacks into the self-complementary d(CGCAGAATTCGCG) duplex in solution. , 1982, Biochemistry.

[40]  Z. Otwinowski,et al.  [20] Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[41]  I. Tinoco,et al.  18 RNA Structural Elements and RNA Function , 1993 .