data_6115 ####################### # Entry information # ####################### save_entry_information _Saveframe_category entry_information _Entry_title ; Structural and Functional Analysis of a picornaviral Internal cis-acting replication element ; _BMRB_accession_number 6115 _BMRB_flat_file_name bmr6115.str _Entry_type original _Submission_date 2004-02-25 _Accession_date 2004-02-25 _Entry_origination author _NMR_STAR_version 2.1.1 _Experimental_method NMR _Details . loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Kaluarachchi Kumaralal . . 2 Thiviyanathan Varatharasa . . stop_ loop_ _Saveframe_category_type _Saveframe_category_type_count assigned_chemical_shifts 1 stop_ loop_ _Data_type _Data_type_count "1H chemical shifts" 268 stop_ loop_ _Revision_date _Revision_keyword _Revision_author _Revision_detail 2004-09-28 original author . stop_ save_ ############################# # Citation for this entry # ############################# save_entry_citation _Saveframe_category entry_citation _Citation_full . _Citation_title 'High resolution Structure of a Picornaviral Internal Cis-acting RNA Replication Element (cre)' _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 15314212 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Thiviyanathan Varatharasa . . 2 Yang Y. . . 3 Kaluarachchi Kumaralal . . 4 Rijnbrand R. . . 5 Gorenstein D. G. . 6 Lemon S. M. . stop_ _Journal_abbreviation 'Proc. Natl. Acad. Sci. U.S.A.' _Journal_volume 101 _Journal_issue 34 _Journal_CSD . _Book_chapter_title . _Book_volume . _Book_series . _Book_ISBN . _Conference_state_province . _Conference_abstract_number . _Page_first 12688 _Page_last 12693 _Year 2004 _Details . loop_ _Keyword rhinovirus cre-structure stem-loop uridylation stop_ save_ ####################################### # Cited references within the entry # ####################################### save_ref_1 _Saveframe_category citation _Citation_full ; Yang Y, Rijnbrand R, McKnight KL, Wimmer E, Paul A, Martin A, Lemon SM. J Virol. 2002 Aug;76(15):7485-94. ; _Citation_title 'Sequence requirements for viral RNA replication and VPg uridylylation directed by the internal cis-acting replication element (cre) of human rhinovirus type 14.' _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 12097561 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Yang Yan . . 2 Rijnbrand Rene . . 3 McKnight 'Kevin L.' L. . 4 Wimmer Eckard . . 5 Paul Aniko . . 6 Martin Annette . . 7 Lemon 'Stanley M.' M. . stop_ _Journal_abbreviation 'J. Virol.' _Journal_name_full 'Journal of virology' _Journal_volume 76 _Journal_issue 15 _Journal_CSD . _Book_title . _Book_chapter_title . _Book_volume . _Book_series . _Book_publisher . _Book_publisher_city . _Book_ISBN . _Conference_title . _Conference_site . _Conference_state_province . _Conference_country . _Conference_start_date . _Conference_end_date . _Conference_abstract_number . _Thesis_institution . _Thesis_institution_city . _Thesis_institution_country . _Page_first 7485 _Page_last 7494 _Year 2002 _Details ; Until recently, the cis-acting signals required for replication of picornaviral RNAs were believed to be restricted to the 5' and 3' noncoding regions of the genome. However, an RNA stem-loop in the VP1-coding sequence of human rhinovirus type 14 (HRV-14) is essential for viral minus-strand RNA synthesis (K. L. McKnight and S. M. Lemon, RNA 4:1569-1584, 1998). The nucleotide sequence of the apical loop of this internal cis-acting replication element (cre) was critical for RNA synthesis, while secondary RNA structure, but not primary sequence, was shown to be important within the duplex stem. Similar cres have since been identified in other picornaviral genomes. These RNA segments appear to serve as template for the uridylylation of the genome-linked protein, VPg, providing the VPg-pUpU primer required for viral RNA transcription (A. V. Paul et al., J. Virol. 74:10359-10370, 2000). Here, we show that the minimal functional HRV-14 cre resides within a 33-nucleotide (nt) RNA segment that is predicted to form a simple stem-loop with a 14-nt loop sequence. An extensive mutational analysis involving every possible base substitution at each position within the loop segment defined the sequence that is required within this loop for efficient replication of subgenomic HRV-14 replicon RNAs. These results indicate that three consecutive adenosine residues (nt 2367 to 2369) within the 5' half of this loop are critically important for cre function and suggest that a common RNNNAARNNNNNNR loop motif exists among the cre sequences of enteroviruses and rhinoviruses. We found a direct, positive correlation between the capacity of mutated cres to support RNA replication and their ability to function as template in an in vitro VPg uridylylation reaction, suggesting that these functions are intimately linked. These data thus define more precisely the sequence and structural requirements of the HRV-14 cre and provide additional support for a model in which the role of the cre in RNA replication is to act as template for VPg uridylylation. ; save_ save_ref_2 _Saveframe_category citation _Citation_full ; McKnight KL, Lemon SM. RNA. 1998 Dec;4(12):1569-84. ; _Citation_title 'The rhinovirus type 14 genome contains an internally located RNA structure that is required for viral replication.' _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 9848654 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 McKnight 'K. L.' L. . 2 Lemon 'S. M.' M. . stop_ _Journal_abbreviation RNA _Journal_name_full 'RNA (New York, N.Y.)' _Journal_volume 4 _Journal_issue 12 _Journal_CSD . _Book_title . _Book_chapter_title . _Book_volume . _Book_series . _Book_publisher . _Book_publisher_city . _Book_ISBN . _Conference_title . _Conference_site . _Conference_state_province . _Conference_country . _Conference_start_date . _Conference_end_date . _Conference_abstract_number . _Thesis_institution . _Thesis_institution_city . _Thesis_institution_country . _Page_first 1569 _Page_last 1584 _Year 1998 _Details ; Cis-acting RNA signals are required for replication of positive-strand viruses such as the picornaviruses. Although these generally have been mapped to the 5' and/or 3' termini of the viral genome, RNAs derived from human rhinovirus type 14 are unable to replicate unless they contain an internal cis-acting replication element (cre) located within the genome segment encoding the capsid proteins. Here, we show that the essential cre sequence is 83-96 nt in length and located between nt 2318-2413 of the genome. Using dicistronic RNAs in which translation of the P1 and P2-P3 segments of the polyprotein were functionally dissociated, we further demonstrate that translation of the cre sequence is not required for RNA replication. Thus, although it is located within a protein-coding segment of the genome, the cre functions as an RNA entity. Computer folds suggested that cre sequences could form a stable structure in either positive- or minus-strand RNA. However, an analysis of mutant RNAs containing multiple covariant and non-covariant nucleotide substitutions within these putative structures demonstrated that only the predicted positive-strand structure is essential for efficient RNA replication. The absence of detectable minus-strand synthesis from RNAs that lack the cre suggests that the cre is required for initiation of minus-strand RNA synthesis. Since a lethal 3' noncoding region mutation could be partially rescued by a compensating mutation within the cre, the cre appears to participate in a long-range RNA-RNA interaction required for this process. These data provide novel insight into the mechanisms of replication of a positive-strand RNA virus, as they define the involvement of an internally located RNA structure in the recognition of viral RNA by the viral replicase complex. Since internally located RNA replication signals have been shown to exist in several other positive-strand RNA virus families, these observations are potentially relevant to a wide array of related viruses. ; save_ ################################## # Molecular system description # ################################## save_system_CRE _Saveframe_category molecular_system _Mol_system_name 'Rhonivirus CRE' _Abbreviation_common CRE _Enzyme_commission_number . loop_ _Mol_system_component_name _Mol_label cre $CRE stop_ _System_molecular_weight . _System_physical_state native _System_oligomer_state monomer _System_paramagnetic no _System_thiol_state 'not present' _Database_query_date . _Details . save_ ######################## # Monomeric polymers # ######################## save_CRE _Saveframe_category monomeric_polymer _Mol_type polymer _Mol_polymer_class RNA _Name_common 'Cis-acting replication element' _Abbreviation_common CRE _Molecular_mass . _Mol_thiol_state 'not present' _Details . ############################## # Polymer residue sequence # ############################## _Residue_count 34 _Mol_residue_sequence ; GGUCAUCGUUGAGAAAACGA AACAGACGGUGGCC ; loop_ _Residue_seq_code _Residue_author_seq_code _Residue_label 1 1 G 2 2 G 3 3 U 4 4 C 5 5 A 6 6 U 7 7 C 8 8 G 9 9 U 10 10 U 11 11 G 12 12 A 13 13 G 14 14 A 15 15 A 16 16 A 17 17 A 18 18 C 19 19 G 20 20 A 21 21 A 22 22 A 23 23 C 24 24 A 25 25 G 26 26 A 27 27 C 28 28 G 29 29 G 30 30 U 31 31 G 32 32 G 33 33 C 34 34 C stop_ _Sequence_homology_query_date . _Sequence_homology_query_revised_last_date . save_ #################### # Natural source # #################### save_natural_source _Saveframe_category natural_source loop_ _Mol_label _Organism_name_common _NCBI_taxonomy_ID _Superkingdom _Kingdom _Genus _Species $CRE . . viruses . . . stop_ save_ ######################### # Experimental source # ######################### save_experimental_source _Saveframe_category experimental_source loop_ _Mol_label _Production_method _Host_organism_name_common _Genus _Species _Strain _Vector_name _Details $CRE 'enzymatic semisynthesis' . . . . . 'In-vitro synthesis of DNA templates using T7 RNA polymerase' stop_ save_ ##################################### # Sample contents and methodology # ##################################### ######################## # Sample description # ######################## save_sample_1 _Saveframe_category sample _Sample_type solution _Details . loop_ _Mol_label _Concentration_value _Concentration_value_units _Concentration_min_value _Concentration_max_value _Isotopic_labeling $CRE . mM 0.8 2.0 '[U-13C; U-15N]' stop_ save_ ############################ # Computer software used # ############################ save_FELIX _Saveframe_category software _Name FELIX _Version 2000 loop_ _Task 'peak assignmnets' 'volume measurements' stop_ _Details . save_ ######################### # Experimental detail # ######################### ################################## # NMR Spectrometer definitions # ################################## save_NMR_spectrometer_1 _Saveframe_category NMR_spectrometer _Manufacturer Varian _Model UnityPlus _Field_strength 600 _Details . save_ save_NMR_spectrometer_2 _Saveframe_category NMR_spectrometer _Manufacturer Varian _Model UnityPlus _Field_strength 750 _Details . save_ ############################# # NMR applied experiments # ############################# save_Seperate_1H-15N_HSQC_experiments_were_done,_optimized_for_imino_and_amino_1 _Saveframe_category NMR_applied_experiment _Experiment_name 'Seperate 1H-15N HSQC experiments were done, optimized for imino and amino' _Sample_label $sample_1 save_ save_protons_2 _Saveframe_category NMR_applied_experiment _Experiment_name protons _Sample_label $sample_1 save_ ####################### # Sample conditions # ####################### save_ex-cond_1 _Saveframe_category sample_conditions _Details . loop_ _Variable_type _Variable_value _Variable_value_error _Variable_value_units pH 6.8 0.1 n/a temperature 298 1 K stop_ save_ #################### # NMR parameters # #################### ############################## # Assigned chemical shifts # ############################## ################################ # Chemical shift referencing # ################################ save_chemical_shift_reference _Saveframe_category chemical_shift_reference _Details . loop_ _Mol_common_name _Atom_type _Atom_isotope_number _Atom_group _Chem_shift_units _Chem_shift_value _Reference_method _Reference_type _External_reference_sample_geometry _External_reference_location _External_reference_axis _Indirect_shift_ratio DSS H 1 'methyl protons' ppm 0.0 internal direct . . . 1.0 stop_ save_ ################################### # Assigned chemical shift lists # ################################### ################################################################### # Chemical Shift Ambiguity Index Value Definitions # # # # The values other than 1 are used for those atoms with different # # chemical shifts that cannot be assigned to stereospecific atoms # # or to specific residues or chains. # # # # Index Value Definition # # # # 1 Unique (including isolated methyl protons, # # geminal atoms, and geminal methyl # # groups with identical chemical shifts) # # (e.g. ILE HD11, HD12, HD13 protons) # # 2 Ambiguity of geminal atoms or geminal methyl # # proton groups (e.g. ASP HB2 and HB3 # # protons, LEU CD1 and CD2 carbons, or # # LEU HD11, HD12, HD13 and HD21, HD22, # # HD23 methyl protons) # # 3 Aromatic atoms on opposite sides of # # symmetrical rings (e.g. TYR HE1 and HE2 # # protons) # # 4 Intraresidue ambiguities (e.g. LYS HG and # # HD protons or TRP HZ2 and HZ3 protons) # # 5 Interresidue ambiguities (LYS 12 vs. LYS 27) # # 6 Intermolecular ambiguities (e.g. ASP 31 CA # # in monomer 1 and ASP 31 CA in monomer 2 # # of an asymmetrical homodimer, duplex # # DNA assignments, or other assignments # # that may apply to atoms in one or more # # molecule in the molecular assembly) # # 9 Ambiguous, specific ambiguity not defined # # # ################################################################### save_shift1 _Saveframe_category assigned_chemical_shifts _Details . loop_ _Sample_label $sample_1 stop_ _Sample_conditions_label $ex-cond_1 _Chem_shift_reference_set_label $chemical_shift_reference _Mol_system_component_name cre _Text_data_format . _Text_data . loop_ _Atom_shift_assign_ID _Residue_author_seq_code _Residue_seq_code _Residue_label _Atom_name _Atom_type _Chem_shift_value _Chem_shift_value_error _Chem_shift_ambiguity_code 1 1 1 G H1 H 13.45 0.01 1 2 1 1 G H8 H 8.15 0.01 1 3 1 1 G H1' H 5.86 0.01 1 4 1 1 G H2' H 4.98 0.01 1 5 1 1 G H3' H 4.73 0.01 1 6 1 1 G H4' H 4.44 0.01 1 7 1 1 G H5' H 4.01 0.01 1 8 1 1 G H5'' H 3.94 0.01 1 9 2 2 G H1 H 13.57 0.01 1 10 2 2 G H8 H 7.62 0.01 1 11 2 2 G H1' H 5.97 0.01 1 12 2 2 G H2' H 4.60 0.01 1 13 2 2 G H3' H 4.47 0.01 1 14 2 2 G H4' H 4.42 0.01 1 15 2 2 G H5' H 4.42 0.01 1 16 2 2 G H5'' H 4.06 0.01 1 17 3 3 U H3 H 12.40 0.01 1 18 3 3 U H5 H 5.48 0.01 1 19 3 3 U H6 H 7.84 0.01 1 20 3 3 U H1' H 5.58 0.01 1 21 3 3 U H2' H 4.21 0.01 1 22 3 3 U H3' H 4.47 0.01 1 23 3 3 U H4' H 4.39 0.01 1 24 3 3 U H5' H 4.41 0.01 1 25 3 3 U H5'' H 4.08 0.01 1 26 4 4 C H41 H 8.45 0.01 2 27 4 4 C H42 H 7.03 0.01 2 28 4 4 C H5 H 5.73 0.01 1 29 4 4 C H6 H 7.99 0.01 1 30 4 4 C H1' H 5.69 0.01 1 31 4 4 C H2' H 4.44 0.01 1 32 4 4 C H3' H 4.26 0.01 1 33 4 4 C H4' H 4.20 0.01 1 34 4 4 C H5' H 4.14 0.01 1 35 4 4 C H5'' H 4.05 0.01 1 36 5 5 A H2 H 7.40 0.01 1 37 5 5 A H61 H 6.59 0.01 1 38 5 5 A H62 H 8.03 0.01 1 39 5 5 A H8 H 8.01 0.01 1 40 5 5 A H1' H 5.87 0.01 1 41 5 5 A H2' H 4.62 0.01 1 42 5 5 A H3' H 4.47 0.01 1 43 5 5 A H4' H 0.36 0.01 1 44 5 5 A H5' H 4.28 0.01 1 45 5 5 A H5'' H 4.17 0.01 1 46 6 6 U H3 H 12.06 0.01 1 47 6 6 U H5 H 5.37 0.01 1 48 6 6 U H6 H 7.69 0.01 1 49 6 6 U H1' H 5.34 0.01 1 50 6 6 U H2' H 4.67 0.01 1 51 6 6 U H3' H 4.52 0.01 1 52 6 6 U H4' H 4.27 0.01 1 53 7 7 C H41 H 8.36 0.01 2 54 7 7 C H42 H 6.84 0.01 2 55 7 7 C H5 H 5.64 0.01 1 56 7 7 C H6 H 7.93 0.01 1 57 7 7 C H1' H 5.67 0.01 1 58 7 7 C H2' H 4.59 0.01 1 59 7 7 C H3' H 4.42 0.01 1 60 7 7 C H4' H 4.31 0.01 1 61 7 7 C H5' H 4.13 0.01 1 62 7 7 C H5'' H 4.09 0.01 1 63 8 8 G H1 H 12.90 0.01 1 64 8 8 G H21 H 6.09 0.01 2 65 8 8 G H8 H 7.91 0.01 1 66 8 8 G H1' H 5.79 0.01 1 67 8 8 G H2' H 4.63 0.01 1 68 8 8 G H3' H 4.34 0.01 1 69 8 8 G H4' H 4.20 0.01 1 70 9 9 U H3 H 14.18 0.01 1 71 9 9 U H5 H 5.75 0.01 1 72 9 9 U H6 H 7.81 0.01 1 73 9 9 U H1' H 5.58 0.01 1 74 9 9 U H2' H 4.64 0.01 1 75 9 9 U H3' H 4.47 0.01 1 76 9 9 U H4' H 4.32 0.01 1 77 9 9 U H5' H 4.19 0.01 1 78 9 9 U H5'' H 4.05 0.01 1 79 10 10 U H3 H 11.60 0.01 1 80 10 10 U H5 H 5.02 0.01 1 81 10 10 U H6 H 7.68 0.01 1 82 10 10 U H1' H 5.67 0.01 1 83 10 10 U H2' H 4.43 0.01 1 84 10 10 U H3' H 4.39 0.01 1 85 10 10 U H4' H 4.44 0.01 1 86 11 11 G H1 H 13.02 0.01 1 87 11 11 G H8 H 7.52 0.01 1 88 11 11 G H1' H 5.63 0.01 1 89 11 11 G H2' H 4.59 0.01 1 90 11 11 G H3' H 4.41 0.01 1 91 11 11 G H4' H 4.31 0.01 1 92 11 11 G H5' H 4.13 0.01 1 93 12 12 A H2 H 7.84 0.01 1 94 12 12 A H8 H 7.94 0.01 1 95 13 13 G H8 H 7.67 0.01 1 96 13 13 G H1' H 5.48 0.01 1 97 14 14 A H2 H 7.88 0.01 1 98 14 14 A H8 H 8.12 0.01 1 99 14 14 A H1' H 5.79 0.01 1 100 14 14 A H2' H 4.45 0.01 1 101 14 14 A H3' H 4.42 0.01 1 102 14 14 A H4' H 4.31 0.01 1 103 14 14 A H5' H 4.12 0.01 1 104 14 14 A H5'' H 4.10 0.01 1 105 15 15 A H2 H 7.82 0.01 1 106 15 15 A H8 H 8.12 0.01 1 107 15 15 A H1' H 5.83 0.01 1 108 15 15 A H2' H 4.64 0.01 1 109 15 15 A H3' H 4.56 0.01 1 110 15 15 A H4' H 4.46 0.01 1 111 15 15 A H5' H 4.14 0.01 1 112 15 15 A H5'' H 4.08 0.01 1 113 16 16 A H2 H 7.76 0.01 1 114 16 16 A H8 H 7.87 0.01 1 115 16 16 A H1' H 5.58 0.01 1 116 16 16 A H2' H 4.48 0.01 1 117 16 16 A H3' H 4.39 0.01 1 118 16 16 A H4' H 4.15 0.01 1 119 16 16 A H5' H 4.11 0.01 1 120 16 16 A H5'' H 4.11 0.01 1 121 17 17 A H2 H 7.90 0.01 1 122 17 17 A H8 H 7.93 0.01 1 123 17 17 A H1' H 5.51 0.01 1 124 17 17 A H2' H 4.41 0.01 1 125 17 17 A H3' H 4.67 0.01 1 126 17 17 A H4' H 4.40 0.01 1 127 17 17 A H5' H 4.14 0.01 1 128 17 17 A H5'' H 4.01 0.01 1 129 18 18 C H5 H 5.55 0.01 1 130 18 18 C H6 H 7.72 0.01 1 131 18 18 C H1' H 5.45 0.01 1 132 18 18 C H2' H 4.40 0.01 1 133 18 18 C H3' H 4.42 0.01 1 134 19 19 G H8 H 7.77 0.01 1 135 19 19 G H1' H 5.56 0.01 1 136 19 19 G H2' H 4.42 0.01 1 137 19 19 G H3' H 4.35 0.01 1 138 19 19 G H4' H 4.19 0.01 1 139 20 20 A H2 H 7.98 0.01 1 140 20 20 A H8 H 8.14 0.01 1 141 20 20 A H1' H 5.84 0.01 1 142 20 20 A H2' H 4.49 0.01 1 143 20 20 A H3' H 4.28 0.01 1 144 20 20 A H4' H 4.18 0.01 1 145 20 20 A H5' H 4.12 0.01 1 146 20 20 A H5'' H 4.05 0.01 1 147 21 21 A H2 H 7.78 0.01 1 148 21 21 A H8 H 8.08 0.01 1 149 21 21 A H1' H 5.79 0.01 1 150 21 21 A H2' H 4.64 0.01 1 151 21 21 A H3' H 4.46 0.01 1 152 21 21 A H4' H 4.45 0.01 1 153 21 21 A H5' H 4.47 0.01 1 154 21 21 A H5'' H 4.06 0.01 1 155 22 22 A H2 H 7.86 0.01 1 156 22 22 A H8 H 8.04 0.01 1 157 22 22 A H1' H 5.79 0.01 1 158 22 22 A H2' H 4.34 0.01 1 159 22 22 A H4' H 4.31 0.01 1 160 22 22 A H5' H 4.14 0.01 1 161 22 22 A H5'' H 4.00 0.01 1 162 23 23 C H5 H 5.57 0.01 1 163 23 23 C H6 H 7.57 0.01 1 164 23 23 C H1' H 5.66 0.01 1 165 23 23 C H2' H 4.33 0.01 1 166 23 23 C H3' H 4.60 0.01 1 167 23 23 C H4' H 4.26 0.01 1 168 23 23 C H5' H 4.12 0.01 1 169 23 23 C H5'' H 4.12 0.01 1 170 24 24 A H2 H 7.87 0.01 1 171 24 24 A H8 H 8.17 0.01 1 172 24 24 A H1' H 6.01 0.01 1 173 24 24 A H2' H 4.62 0.01 1 174 24 24 A H3' H 4.67 0.01 1 175 24 24 A H4' H 4.39 0.01 1 176 24 24 A H5' H 4.21 0.01 1 177 24 24 A H5'' H 4.05 0.01 1 178 25 25 G H1 H 10.51 0.01 1 179 25 25 G H21 H 6.30 0.01 2 180 25 25 G H22 H 6.30 0.01 2 181 25 25 G H8 H 7.72 0.01 1 182 25 25 G H1' H 5.33 0.01 1 183 25 25 G H2' H 4.67 0.01 1 184 25 25 G H3' H 4.49 0.01 1 185 25 25 G H4' H 4.23 0.01 1 186 25 25 G H5' H 4.06 0.01 1 187 26 26 A H2 H 7.66 0.01 1 188 26 26 A H61 H 5.93 0.01 2 189 26 26 A H62 H 5.50 0.01 2 190 26 26 A H8 H 7.80 0.01 1 191 26 26 A H1' H 5.93 0.01 1 192 26 26 A H2' H 4.56 0.01 1 193 26 26 A H3' H 4.61 0.01 1 194 26 26 A H4' H 4.52 0.01 1 195 26 26 A H5' H 4.11 0.01 1 196 26 26 A H5'' H 4.11 0.01 1 197 27 27 C H41 H 8.27 0.01 1 198 27 27 C H42 H 6.97 0.01 1 199 27 27 C H5 H 5.18 0.01 1 200 27 27 C H6 H 7.40 0.01 1 201 27 27 C H1' H 5.46 0.01 1 202 27 27 C H2' H 4.54 0.01 1 203 27 27 C H3' H 4.59 0.01 1 204 27 27 C H4' H 4.43 0.01 1 205 27 27 C H5' H 4.06 0.01 1 206 28 28 G H1 H 12.36 0.01 1 207 28 28 G H21 H 8.58 0.01 2 208 28 28 G H8 H 7.58 0.01 1 209 28 28 G H1' H 5.81 0.01 1 210 28 28 G H2' H 4.60 0.01 1 211 28 28 G H3' H 4.52 0.01 1 212 28 28 G H4' H 4.29 0.01 1 213 28 28 G H5' H 4.14 0.01 1 214 28 28 G H5'' H 4.08 0.01 1 215 29 29 G H1 H 11.56 0.01 1 216 29 29 G H21 H 8.50 0.01 2 217 29 29 G H8 H 7.24 0.01 1 218 29 29 G H1' H 5.83 0.01 1 219 29 29 G H2' H 4.50 0.01 1 220 29 29 G H3' H 4.34 0.01 1 221 29 29 G H4' H 4.28 0.01 1 222 29 29 G H5' H 4.11 0.01 1 223 29 29 G H5'' H 4.10 0.01 1 224 30 30 U H3 H 13.54 0.01 1 225 30 30 U H5 H 5.29 0.01 1 226 30 30 U H6 H 7.69 0.01 1 227 30 30 U H1' H 5.59 0.01 1 228 30 30 U H2' H 4.60 0.01 1 229 30 30 U H3' H 4.55 0.01 1 230 30 30 U H4' H 4.31 0.01 1 231 30 30 U H5' H 4.19 0.01 2 232 30 30 U H5'' H 4.05 0.01 2 233 31 31 G H1 H 12.10 0.01 1 234 31 31 G H21 H 5.85 0.01 2 235 31 31 G H8 H 7.78 0.01 1 236 31 31 G H1' H 5.88 0.01 1 237 31 31 G H2' H 4.59 0.01 1 238 31 31 G H3' H 4.94 0.01 1 239 31 31 G H4' H 4.44 0.01 1 240 31 31 G H5' H 4.23 0.01 1 241 31 31 G H5'' H 4.14 0.01 1 242 32 32 G H1 H 11.51 0.01 1 243 32 32 G H21 H 6.40 0.01 2 244 32 32 G H22 H 6.40 0.01 2 245 32 32 G H8 H 7.35 0.01 1 246 32 32 G H1' H 5.81 0.01 1 247 32 32 G H2' H 4.54 0.01 1 248 32 32 G H3' H 4.70 0.01 1 249 32 32 G H4' H 4.38 0.01 1 250 32 32 G H5' H 4.21 0.01 1 251 32 32 G H5'' H 4.05 0.01 1 252 33 33 C H41 H 8.49 0.01 2 253 33 33 C H42 H 7.01 0.01 2 254 33 33 C H5 H 5.40 0.01 1 255 33 33 C H6 H 7.72 0.01 1 256 33 33 C H1' H 5.83 0.01 1 257 33 33 C H2' H 4.64 0.01 1 258 33 33 C H3' H 4.51 0.01 1 259 33 33 C H4' H 4.14 0.01 1 260 33 33 C H5' H 4.05 0.01 1 261 34 34 C H5 H 5.59 0.01 1 262 34 34 C H6 H 7.55 0.01 1 263 34 34 C H1' H 5.69 0.01 1 264 34 34 C H2' H 4.18 0.01 1 265 34 34 C H3' H 4.29 0.01 1 266 34 34 C H4' H 4.16 0.01 1 267 34 34 C H5' H 4.25 0.01 1 268 34 34 C H5'' H 4.11 0.01 1 stop_ save_