data_7090 ####################### # Entry information # ####################### save_entry_information _Saveframe_category entry_information _Entry_title ; Solution structure of a purine rich hexaloop hairpin belonging to PGY/MDR1 mRNA and targeted by antisense oligonucleotides ; _BMRB_accession_number 7090 _BMRB_flat_file_name bmr7090.str _Entry_type original _Submission_date 2006-04-27 _Accession_date 2006-04-27 _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 Joli Flore . . 2 Bouchemal Nadia . . 3 Laigle Alain . . 4 Hartmann Brigitte . . 5 Hantz Edith . . stop_ loop_ _Saveframe_category_type _Saveframe_category_type_count assigned_chemical_shifts 1 stop_ loop_ _Data_type _Data_type_count "1H chemical shifts" 149 "13C chemical shifts" 90 "15N chemical shifts" 21 "31P chemical shifts" 17 stop_ loop_ _Revision_date _Revision_keyword _Revision_author _Revision_detail 2007-04-27 original author . stop_ _Original_release_date 2007-04-27 save_ ############################# # Citation for this entry # ############################# save_entry_citation _Saveframe_category entry_citation _Citation_full . _Citation_title ; Solution structure of a purine rich hexaloop hairpin belonging to PGY/MDR1 mRNA and targeted by antisense oligonucleotides ; _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 17041234 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Joli Flore . . 2 Bouchemal Nadia . . 3 Laigle Alain . . 4 Hartmann Brigitte . . 5 Hantz Edith . . stop_ _Journal_abbreviation 'Nucleic Acids Res.' _Journal_volume 34 _Journal_issue 20 _Journal_CSD . _Book_chapter_title . _Book_volume . _Book_series . _Book_ISBN . _Conference_state_province . _Conference_abstract_number . _Page_first 5740 _Page_last 5751 _Year 2006 _Details . loop_ _Keyword '13C/15N-labeled RNA' 'G.U wobble pair' 'hairpin structure' hexaloop 'RNA structure' stop_ save_ ################################## # Molecular system description # ################################## save_assembly _Saveframe_category molecular_system _Mol_system_name '18 mer hairpin hexaloop' _Enzyme_commission_number . loop_ _Mol_system_component_name _Mol_label '18 mer hairpin hexaloop' $ARNm_MDR1_PGY stop_ _System_molecular_weight . _System_physical_state native _System_oligomer_state 'single stranded RNA' _System_paramagnetic no _System_thiol_state 'not present' loop_ _Magnetic_equivalence_ID _Magnetically_equivalent_system_component 1 '18 mer hairpin hexaloop' stop_ loop_ _Biological_function 'antisens target' 'coding for P-gp' stop_ _Database_query_date . _Details . save_ ######################## # Monomeric polymers # ######################## save_ARNm_MDR1_PGY _Saveframe_category monomeric_polymer _Mol_type polymer _Mol_polymer_class RNA _Name_common '18mer hairpin hexaloop' _Molecular_mass . _Mol_thiol_state 'not present' _Details . ############################## # Polymer residue sequence # ############################## _Residue_count 18 _Mol_residue_sequence GAGGUCGGGAUGGAUCUC loop_ _Residue_seq_code _Residue_label 1 G 2 A 3 G 4 G 5 U 6 C 7 G 8 G 9 G 10 A 11 U 12 G 13 G 14 A 15 U 16 C 17 U 18 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 $ARNm_MDR1_PGY Human 9606 Eukaryota Metazoa Homo sapiens 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 $ARNm_MDR1_PGY 'obtained from a vendor' . . . . . 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 _Isotopic_labeling $ARNm_MDR1_PGY 1 mM '[U-98% 13C; U-98% 15N]' 'phosphate buffer' 10 mM . EDTA 0.01 mM . stop_ save_ ######################### # Experimental detail # ######################### ################################## # NMR Spectrometer definitions # ################################## save_500MHz_spectrometer _Saveframe_category NMR_spectrometer _Manufacturer Varian _Model UnityInova _Field_strength 500 _Details 'Two probes: 5mm gradient indirect detection, 5mm triple (1H, 13C, 15N) detection.' save_ ############################# # NMR applied experiments # ############################# save_1H15N_HSQC_1 _Saveframe_category NMR_applied_experiment _Experiment_name '1H15N HSQC' _Sample_label $sample_1 save_ save_1H13C_HSQC_2 _Saveframe_category NMR_applied_experiment _Experiment_name '1H13C HSQC' _Sample_label $sample_1 save_ save_1H1H13C_HCCH_COSY_3 _Saveframe_category NMR_applied_experiment _Experiment_name '1H1H13C HCCH COSY' _Sample_label $sample_1 save_ save_1H1H13C_NOESY_HSQC_4 _Saveframe_category NMR_applied_experiment _Experiment_name '1H1H13C NOESY HSQC' _Sample_label $sample_1 save_ save_HNN_COSY_5 _Saveframe_category NMR_applied_experiment _Experiment_name 'HNN COSY' _Sample_label $sample_1 save_ save_1H1H13C_HSCQ_TOCSY_6 _Saveframe_category NMR_applied_experiment _Experiment_name '1H1H13C HSCQ TOCSY' _Sample_label $sample_1 save_ save_HCN_TROSY_7 _Saveframe_category NMR_applied_experiment _Experiment_name 'HCN TROSY' _Sample_label $sample_1 save_ save_CU_HNCCCH_8 _Saveframe_category NMR_applied_experiment _Experiment_name 'CU HNCCCH' _Sample_label $sample_1 save_ save_GA_HNC_TOCSY_CH_9 _Saveframe_category NMR_applied_experiment _Experiment_name 'GA HNC TOCSY CH' _Sample_label $sample_1 save_ ####################### # Sample conditions # ####################### save_conditions_1 _Saveframe_category sample_conditions _Details ; pH=6.8 phosphate buffer 10mM EDTA 10^(-5) ; loop_ _Variable_type _Variable_value _Variable_value_error _Variable_value_units 'ionic strength' 10 0.2 mM pH 6.8 0.1 pH stop_ save_ #################### # NMR parameters # #################### ############################## # Assigned chemical shifts # ############################## ################################ # Chemical shift referencing # ################################ save_chemical_shift_referencing _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 methanol C 13 'methyl carbon' ppm 49.30 external indirect . . . 1.0 TSP H 1 protons ppm 0.00 internal direct . . . 1.0 'ammonium chloride' N 15 nitrogen ppm 24.93 external indirect . . . 1.0 'phosphoric acid' P 31 phosphorus ppm 0.00 external indirect . . . 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_chem_shift_list_1 _Saveframe_category assigned_chemical_shifts _Details . loop_ _Experiment_label '1H15N HSQC' '1H13C HSQC' '1H1H13C HCCH COSY' '1H1H13C NOESY HSQC' 'HNN COSY' '1H1H13C HSCQ TOCSY' 'HCN TROSY' 'CU HNCCCH' 'GA HNC TOCSY CH' stop_ loop_ _Sample_label $sample_1 stop_ _Sample_conditions_label $conditions_1 _Chem_shift_reference_set_label $chemical_shift_referencing _Mol_system_component_name '18 mer hairpin hexaloop' _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 12.31 0.01 1 2 1 1 G H8 H 7.97 0.01 1 3 1 1 G N1 N 146.38 0.01 1 4 1 1 G N9 N 168.10 0.01 1 5 1 1 G H1' H 5.62 0.01 1 6 1 1 G H2' H 4.77 0.01 1 7 1 1 G H3' H 4.53 0.01 1 8 1 1 G H4' H 4.03 0.01 1 9 1 1 G H5' H 4.22 0.01 2 10 1 1 G H5'' H 4.42 0.01 2 11 1 1 G C1' C 93.78 0.01 1 12 1 1 G C2' C 74.68 0.01 1 13 1 1 G C3' C 73.77 0.01 1 14 1 1 G C4' C 83.59 0.01 1 15 1 1 G C5' C 66.81 0.01 1 16 1 1 G P P -0.19 0.01 1 17 2 2 A H2 H 7.63 0.01 1 18 2 2 A H8 H 8.05 0.01 1 19 2 2 A N9 N 167.78 0.01 1 20 2 2 A H1' H 6.15 0.01 1 21 2 2 A H2' H 4.75 0.01 1 22 2 2 A H3' H 4.57 0.01 1 23 2 2 A H4' H 4.71 0.01 1 24 2 2 A H5' H 4.52 0.01 2 25 2 2 A H5'' H 4.12 0.01 2 26 2 2 A C1' C 93.00 0.01 1 27 2 2 A C2' C 7.81 0.01 1 28 2 2 A C3' C 72.48 0.01 1 29 2 2 A C4' C 82.60 0.01 1 30 2 2 A C5' C 64.95 0.01 1 31 2 2 A P P -0.45 0.01 1 32 3 3 G H1 H 12.86 0.01 1 33 3 3 G H8 H 7.25 0.01 1 34 3 3 G N1 N 144.97 0.01 1 35 3 3 G N9 N 169.00 0.01 1 36 3 3 G H1' H 5.75 0.01 1 37 3 3 G H2' H 4.53 0.01 1 38 3 3 G H3' H 4.22 0.01 1 39 3 3 G H4' H 4.70 0.01 1 40 3 3 G H5' H 4.19 0.01 2 41 3 3 G H5'' H 4.49 0.01 2 42 3 3 G C1' C 92.30 0.01 1 43 3 3 G C2' C 74.68 0.01 1 44 3 3 G C3' C 73.75 0.01 1 45 3 3 G C4' C 96.10 0.01 1 46 3 3 G C5' C 67.80 0.01 1 47 3 3 G P P -0.45 0.01 1 48 4 4 G H1 H 11.53 0.01 1 49 4 4 G H8 H 7.13 0.01 1 50 4 4 G N1 N 145.28 0.01 1 51 4 4 G H1' H 5.77 0.01 1 52 4 4 G H2' H 4.69 0.01 1 53 4 4 G H3' H 4.46 0.01 1 54 4 4 G H4' H 4.22 0.01 1 55 4 4 G H5' H 4.45 0.01 2 56 4 4 G H5'' H 4.06 0.01 2 57 4 4 G C1' C 93.65 0.01 1 58 4 4 G C2' C 76.87 0.01 1 59 4 4 G C3' C 72.34 0.01 1 60 4 4 G C4' C 83.67 0.01 1 61 4 4 G C5' C 65.41 0.01 1 62 4 4 G P P -0.83 0.01 1 63 5 5 U H3 H 14.38 0.01 1 64 5 5 U H5 H 5.20 0.01 1 65 5 5 U H6 H 7.69 0.01 1 66 5 5 U N1 N 163.22 0.01 1 67 5 5 U N3 N 146.82 0.01 1 68 5 5 U H1' H 5.50 0.01 1 69 5 5 U H2' H 4.58 0.01 1 70 5 5 U H3' H 4.69 0.01 1 71 5 5 U H4' H 4.20 0.01 1 72 5 5 U H5' H 4.05 0.01 2 73 5 5 U H5'' H 4.48 0.01 2 74 5 5 U C1' C 90.86 0.01 1 75 5 5 U C2' C 75.26 0.01 1 76 5 5 U C3' C 73.81 0.01 1 77 5 5 U C4' C 83.61 0.01 1 78 5 5 U C5' C 65.42 0.01 1 79 5 5 U P P -0.03 0.01 1 80 6 6 C H41 H 8.22 0.01 2 81 6 6 C H42 H 6.65 0.01 2 82 6 6 C H5 H 5.51 0.01 1 83 6 6 C H6 H 7.73 0.01 1 84 6 6 C N4 N 97.51 0.01 1 85 6 6 C H1' H 5.57 0.01 1 86 6 6 C H2' H 4.36 0.01 1 87 6 6 C H3' H 4.48 0.01 1 88 6 6 C H4' H 4.84 0.01 1 89 6 6 C H5' H 4.56 0.01 2 90 6 6 C H5'' H 4.17 0.01 2 91 6 6 C C1' C 93.63 0.01 1 92 6 6 C C2' C 75.49 0.01 1 93 6 6 C C3' C 72.48 0.01 1 94 6 6 C C4' C 84.32 0.01 1 95 6 6 C C5' C 64.77 0.01 1 96 6 6 C P P -0.16 0.01 1 97 7 7 G H8 H 7.89 0.01 1 98 7 7 G N9 N 167.73 0.01 1 99 7 7 G H1' H 5.73 0.01 1 100 7 7 G H2' H 4.04 0.01 1 101 7 7 G H3' H 4.23 0.01 1 102 7 7 G H4' H 4.77 0.01 1 103 7 7 G H5' H 4.10 0.01 2 104 7 7 G H5'' H 4.55 0.01 2 105 7 7 G C1' C 92.08 0.01 1 106 7 7 G C2' C 77.58 0.01 1 107 7 7 G C3' C 73.48 0.01 1 108 7 7 G C4' C 83.05 0.01 1 109 7 7 G C5' C 64.73 0.01 1 110 7 7 G P P -0.90 0.01 1 111 8 8 G H8 H 7.67 0.01 1 112 8 8 G H1' H 5.60 0.01 1 113 8 8 G H2' H 4.35 0.01 1 114 8 8 G H3' H 4.76 0.01 1 115 8 8 G H4' H 4.46 0.01 1 116 8 8 G H5' H 4.14 0.01 2 117 8 8 G H5'' H 4.51 0.01 2 118 8 8 G C1' C 91.95 0.01 1 119 8 8 G C2' C 75.52 0.01 1 120 8 8 G C3' C 73.13 0.01 1 121 8 8 G C4' C 82.02 0.01 1 122 8 8 G C5' C 67.42 0.01 1 123 8 8 G P P -0.00 0.01 1 124 9 9 G H8 H 7.80 0.01 1 125 9 9 G N9 N 168.39 0.01 1 126 9 9 G H1' H 5.88 0.01 1 127 9 9 G H2' H 4.88 0.01 1 128 9 9 G H3' H 4.87 0.01 1 129 9 9 G H4' H 4.57 0.01 1 130 9 9 G H5' H 4.36 0.01 2 131 9 9 G H5'' H 4.30 0.01 2 132 9 9 G C1' C 91.89 0.01 1 133 9 9 G C2' C 76.13 0.01 1 134 9 9 G C3' C 72.67 0.01 1 135 9 9 G C4' C 84.43 0.01 1 136 9 9 G C5' C 66.04 0.01 1 137 9 9 G P P 0.30 0.01 1 138 10 10 A H2 H 7.87 0.01 1 139 10 10 A H8 H 7.98 0.01 1 140 10 10 A N9 N 167.11 0.01 1 141 10 10 A H1' H 5.98 0.01 1 142 10 10 A H2' H 4.42 0.01 1 143 10 10 A H3' H 4.68 0.01 1 144 10 10 A H4' H 4.45 0.01 1 145 10 10 A H5' H 4.17 0.01 2 146 10 10 A H5'' H 4.52 0.01 2 147 10 10 A C1' C 92.33 0.01 1 148 10 10 A C2' C 75.71 0.01 1 149 10 10 A C3' C 73.06 0.01 1 150 10 10 A C4' C 85.12 0.01 1 151 10 10 A C5' C 67.77 0.01 1 152 10 10 A P P -0.03 0.01 1 153 11 11 U H5 H 5.85 0.01 1 154 11 11 U H6 H 7.85 0.01 1 155 11 11 U H1' H 5.96 0.01 1 156 11 11 U H2' H 4.88 0.01 1 157 11 11 U H3' H 4.58 0.01 1 158 11 11 U H4' H 4.36 0.01 1 159 11 11 U H5' H 4.08 0.01 2 160 11 11 U H5'' H 4.29 0.01 2 161 11 11 U C1' C 92.60 0.01 1 162 11 11 U C2' C 76.21 0.01 1 163 11 11 U C3' C 72.39 0.01 1 164 11 11 U C4' C 83.7 0.01 1 165 11 11 U C5' C 66.27 0.01 1 166 11 11 U P P 0.03 0.01 1 167 12 12 G H8 H 8.27 0.01 1 168 12 12 G N9 N 168.38 0.01 1 169 12 12 G H1' H 5.54 0.01 1 170 12 12 G H2' H 4.62 0.01 1 171 12 12 G H3' H 4.82 0.01 1 172 12 12 G H4' H 4.50 0.01 1 173 12 12 G H5' H 4.22 0.01 2 174 12 12 G H5'' H 4.12 0.01 2 175 12 12 G C1' C 90.35 0.01 1 176 12 12 G C2' C 75.06 0.01 1 177 12 12 G C3' C 72.44 0.01 1 178 12 12 G C4' C 85.37 0.01 1 179 12 12 G C5' C 65.41 0.01 1 180 12 12 G P P 0.17 0.01 1 181 13 13 G H1 H 12.22 0.01 1 182 13 13 G H8 H 7.73 0.01 1 183 13 13 G N1 N 145.13 0.01 1 184 13 13 G H1' H 5.63 0.01 1 185 13 13 G H2' H 4.32 0.01 1 186 13 13 G H3' H 4.48 0.01 1 187 13 13 G H4' H 4.76 0.01 1 188 13 13 G H5' H 4.59 0.01 2 189 13 13 G H5'' H 4.40 0.01 2 190 13 13 G C1' C 93.05 0.01 1 191 13 13 G C2' C 75.70 0.01 1 192 13 13 G C3' C 72.47 0.01 1 193 13 13 G C4' C 83.05 0.01 1 194 13 13 G C5' C 66.22 0.01 1 195 13 13 G P P -0.77 0.01 1 196 14 14 A H2 H 7.91 0.01 1 197 14 14 A H8 H 7.89 0.01 1 198 14 14 A N9 N 167.75 0.01 1 199 14 14 A H1' H 6.06 0.01 1 200 14 14 A H2' H 4.71 0.01 1 201 14 14 A H3' H 4.57 0.01 1 202 14 14 A H4' H 4.31 0.01 1 203 14 14 A H5' H 4.53 0.01 2 204 14 14 A H5'' H 4.03 0.01 2 205 14 14 A C1' C 93.16 0.01 1 206 14 14 A C2' C 75.58 0.01 1 207 14 14 A C3' C 72.50 0.01 1 208 14 14 A C4' C 84.79 0.01 1 209 14 14 A C5' C 66.81 0.01 1 210 14 14 A P P -0.19 0.01 1 211 15 15 U H3 H 11.91 0.01 1 212 15 15 U H5 H 5.45 0.01 1 213 15 15 U H6 H 7.66 0.01 1 214 15 15 U N1 N 159.32 0.01 1 215 15 15 U N3 N 157.62 0.01 1 216 15 15 U H1' H 5.38 0.01 1 217 15 15 U H2' H 4.13 0.01 1 218 15 15 U H3' H 4.50 0.01 1 219 15 15 U H4' H 4.43 0.01 1 220 15 15 U H5' H 4.12 0.01 2 221 15 15 U H5'' H 4.52 0.01 2 222 15 15 U C1' C 93.12 0.01 1 223 15 15 U C2' C 75.84 0.01 1 224 15 15 U C3' C 72.41 0.01 1 225 15 15 U C4' C 81.94 0.01 1 226 15 15 U C5' C 64.60 0.01 1 227 15 15 U P P -0.54 0.01 1 228 16 16 C H41 H 8.47 0.01 2 229 16 16 C H42 H 6.83 0.01 2 230 16 16 C H5 H 5.69 0.01 1 231 16 16 C H6 H 7.94 0.01 1 232 16 16 C N4 N 97.55 0.01 1 233 16 16 C H1' H 5.70 0.01 1 234 16 16 C H2' H 4.59 0.01 1 235 16 16 C H3' H 4.87 0.01 1 236 16 16 C H4' H 4.56 0.01 1 237 16 16 C H5' H 4.42 0.01 2 238 16 16 C H5'' H 4.29 0.01 2 239 16 16 C C1' C 92.72 0.01 1 240 16 16 C C2' C 74.49 0.01 1 241 16 16 C C3' C 72.49 0.01 1 242 16 16 C C4' C 82.85 0.01 1 243 16 16 C C5' C 66.80 0.01 1 244 16 16 C P P -0.54 0.01 1 245 17 17 U H3 H 1.2 0.01 1 246 17 17 U H5 H 5.47 0.01 1 247 17 17 U H6 H 7.97 0.01 1 248 17 17 U N1 N 159.63 0.01 1 249 17 17 U N3 N 150.90 0.01 1 250 17 17 U H1' H 5.92 0.01 1 251 17 17 U H2' H 4.06 0.01 1 252 17 17 U H3' H 4.32 0.01 1 253 17 17 U H4' H 4.13 0.01 1 254 17 17 U H5' H 4.48 0.01 2 255 17 17 U H5'' H 4.20 0.01 2 256 17 17 U C1' C 92.07 0.01 1 257 17 17 U C2' C 77.95 0.01 1 258 17 17 U C3' C 72.71 0.01 1 259 17 17 U C4' C 83.92 0.01 1 260 17 17 U C5' C 65.40 0.01 1 261 17 17 U P P -0.02 0.01 1 262 18 18 C H41 H 8.29 0.01 2 263 18 18 C H42 H 6.94 0.01 2 264 18 18 C H5 H 5.73 0.01 1 265 18 18 C H6 H 7.81 0.01 1 266 18 18 C N4 N 97.51 0.01 1 267 18 18 C H1' H 5.86 0.01 1 268 18 18 C H2' H 4.02 0.01 1 269 18 18 C H3' H 4.22 0.01 1 270 18 18 C H4' H 4.43 0.01 1 271 18 18 C H5' H 4.54 0.01 2 272 18 18 C H5'' H 4.11 0.01 2 273 18 18 C C1' C 92.57 0.01 1 274 18 18 C C2' C 77.44 0.01 1 275 18 18 C C3' C 73.48 0.01 1 276 18 18 C C4' C 85.01 0.01 1 277 18 18 C C5' C 64.56 0.01 1 stop_ save_