data_4446 ####################### # Entry information # ####################### save_entry_information _Saveframe_category entry_information _Entry_title ; NMR solution structure of a novel thioredoxin from Bacillus acidocaldarius: possible determinants of protein stability. ; _BMRB_accession_number 4446 _BMRB_flat_file_name bmr4446.str _Entry_type original _Submission_date 1999-10-25 _Accession_date 1999-10-26 _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 Nicastro Giuseppe . . 2 'de Chiara' Cesira . . stop_ loop_ _Saveframe_category_type _Saveframe_category_type_count assigned_chemical_shifts 1 stop_ loop_ _Data_type _Data_type_count "1H chemical shifts" 615 stop_ loop_ _Revision_date _Revision_keyword _Revision_author _Revision_detail 2000-03-10 original author . stop_ _Original_release_date 2000-03-10 save_ ############################# # Citation for this entry # ############################# save_entry_citation _Saveframe_category entry_citation _Citation_full . _Citation_title ; NMR solution structure of a novel thioredoxin from Bacillus acidocaldarius: possible determinants of protein stability. ; _Citation_status submitted _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID ? loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Nicastro Giuseppe . . 2 'de Chiara' Cesira . . stop_ _Journal_abbreviation 'Eur. J. Biochem.' _Journal_name_full 'European journal of Biochemistry' _Journal_volume . _Journal_issue . _Journal_CSD . _Book_chapter_title . _Book_volume . _Book_series . _Book_ISBN . _Conference_state_province . _Conference_abstract_number . _Page_first . _Page_last . _Year . _Details . loop_ _Keyword 'electrostatic interactions' 'NMR spectroscopy' 'solution structure' thermostability thioredoxin stop_ save_ ####################################### # Cited references within the entry # ####################################### save_ref_1 _Saveframe_category citation _Citation_full 'Bartolucci et al. (1997)Biochem. J. 328, 277-285' _Citation_title 'Thioredoxin from Bacillus acidocaldarius: characterization, high-level expression in Escherichia coli and molecular modelling.' _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 9359865 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Bartolucci S . . 2 Guagliardi A . . 3 Pedone E . . 4 'De Pascale' D . . 5 Cannio R . . 6 Camardella L . . 7 Rossi M . . 8 Nicastro G . . 9 'de Chiara' C . . 10 Facci P . . 11 Mascetti G . . 12 Nicolini C . . stop_ _Journal_abbreviation 'Biochem. J.' _Journal_name_full 'The Biochemical journal' _Journal_volume '328 ( Pt 1)' _Journal_issue . _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 277 _Page_last 285 _Year 1997 _Details ; The thioredoxin (Trx) from Bacillus acidocaldarius (BacTrx) was purified to homogeneity by anion-exchange chromatography and gel-filtration chromatography, based on its ability to catalyse the dithiothreitol-dependent reduction of bovine insulin disulphides. The protein has a molecular mass of 11577 Da, determined by electrospray mass spectrometry, a pI of 4.2, and its primary structure was obtained by automated Edman degradation after cleavage with trypsin and cyanogen bromide. The sequences of known bacterial Trxs were aligned at the active site: BacTrx has an identity ranging from 45 to 53% with all sequences except that of the unusual Anabaena strain 7120 Trx (37% identity). The gene coding for BacTrx was isolated by a strategy based on PCR gene amplification and cloned in a plasmid downstream of a lac-derived promoter sequence; the recombinant clone was used as the expression vector for Escherichia coli. The expression was optimized by varying both the time of cell growth and the time of exposure to the inducer isopropyl beta-d-thiogalactoside; expressed BacTrx represents approx. 5% of the total cytosolic protein. CD spectra and differential scanning calorimetry measurements demonstrated that BacTrx is endowed with a higher conformational heat stability than the Trx from E. coli. Nanogravimetry experiments showed a lower content of bound water in BacTrx than in E. coli Trx, and a transition temperature approx. 10 degrees C higher for BacTrx. The three-dimensional model of the oxidized form of BacTrx was constructed by a comparative molecular modelling technique, using E. coli Trx and Anabaena strain 7120 Trx as reference proteins. Increased networks of ion-pairs and shorter loops emerged as major features of the BacTrx structure compared with those of the template proteins. The findings are discussed in the light of the current knowledge about molecular determinants of protein stability. ; save_ ################################## # Molecular system description # ################################## save_thioredoxin _Saveframe_category molecular_system _Mol_system_name thioredoxin _Abbreviation_common thioredoxin _Enzyme_commission_number . loop_ _Mol_system_component_name _Mol_label thioredoxin $Bactrx stop_ _System_molecular_weight . _System_physical_state native _System_oligomer_state monomer _System_paramagnetic no _System_thiol_state 'all disulfide bound' loop_ _Biological_function 'disulfide-containing redox protein' stop_ _Database_query_date . _Details . save_ ######################## # Monomeric polymers # ######################## save_Bactrx _Saveframe_category monomeric_polymer _Mol_type polymer _Mol_polymer_class protein _Name_common 'Thioredoxin from Bacillus acidocaldarius' _Abbreviation_common Bactrx _Molecular_mass 11577 _Mol_thiol_state 'all disulfide bound' _Details . ############################## # Polymer residue sequence # ############################## _Residue_count 105 _Mol_residue_sequence ; ATMTLTDANFQQAIQGDKPV LVDFWAAWCGPCRMMAPVLE EFAEAHADKVTVAKLNVDEN PETTSQFGIMSIPTLILFKG GRPVKQLIGYQPKEQLEAQL ADVLQ ; loop_ _Residue_seq_code _Residue_label 1 ALA 2 THR 3 MET 4 THR 5 LEU 6 THR 7 ASP 8 ALA 9 ASN 10 PHE 11 GLN 12 GLN 13 ALA 14 ILE 15 GLN 16 GLY 17 ASP 18 LYS 19 PRO 20 VAL 21 LEU 22 VAL 23 ASP 24 PHE 25 TRP 26 ALA 27 ALA 28 TRP 29 CYS 30 GLY 31 PRO 32 CYS 33 ARG 34 MET 35 MET 36 ALA 37 PRO 38 VAL 39 LEU 40 GLU 41 GLU 42 PHE 43 ALA 44 GLU 45 ALA 46 HIS 47 ALA 48 ASP 49 LYS 50 VAL 51 THR 52 VAL 53 ALA 54 LYS 55 LEU 56 ASN 57 VAL 58 ASP 59 GLU 60 ASN 61 PRO 62 GLU 63 THR 64 THR 65 SER 66 GLN 67 PHE 68 GLY 69 ILE 70 MET 71 SER 72 ILE 73 PRO 74 THR 75 LEU 76 ILE 77 LEU 78 PHE 79 LYS 80 GLY 81 GLY 82 ARG 83 PRO 84 VAL 85 LYS 86 GLN 87 LEU 88 ILE 89 GLY 90 TYR 91 GLN 92 PRO 93 LYS 94 GLU 95 GLN 96 LEU 97 GLU 98 ALA 99 GLN 100 LEU 101 ALA 102 ASP 103 VAL 104 LEU 105 GLN stop_ _Sequence_homology_query_date . _Sequence_homology_query_revised_last_date 2015-07-14 loop_ _Database_name _Database_accession_code _Database_entry_mol_name _Sequence_query_to_submitted_percentage _Sequence_subject_length _Sequence_identity _Sequence_positive _Sequence_homology_expectation_value PDB 1NSW "The Crystal Structure Of The K18g Mutant Of The Thioredoxin From Alicyclobacillus Acidocaldarius" 100.00 105 99.05 99.05 2.46e-69 PDB 1NW2 "The Crystal Structure Of The Mutant R82e Of Thioredoxin From Alicyclobacillus Acidocaldarius" 100.00 105 99.05 99.05 2.11e-69 PDB 1QUW "Solution Structure Of The Thioredoxin From Bacillus Acidocaldarius" 100.00 105 100.00 100.00 2.33e-70 PDB 1RQM "Solution Structure Of The K18gR82E ALICYCLOBACILLUS Acidocaldarius Thioredoxin Mutant" 100.00 105 98.10 98.10 2.07e-68 GB ACV57898 "thioredoxin [Alicyclobacillus acidocaldarius subsp. acidocaldarius DSM 446]" 100.00 106 100.00 100.00 1.83e-70 GB AEJ42820 "thioredoxin [Alicyclobacillus acidocaldarius subsp. acidocaldarius Tc-4-1]" 98.10 103 99.03 100.00 1.65e-68 GB EED08563 "thioredoxin [Alicyclobacillus acidocaldarius LAA1]" 100.00 106 98.10 98.10 1.44e-68 REF WP_008336730 "thioredoxin [Alicyclobacillus acidocaldarius]" 100.00 106 98.10 98.10 1.44e-68 REF WP_012810252 "thioredoxin [Alicyclobacillus acidocaldarius]" 100.00 106 100.00 100.00 1.83e-70 REF WP_041695007 "thioredoxin [Alicyclobacillus acidocaldarius]" 100.00 106 99.05 100.00 6.08e-70 SP P80579 "RecName: Full=Thioredoxin; Short=Trx" 100.00 105 100.00 100.00 2.33e-70 stop_ save_ #################### # Natural source # #################### save_natural_source _Saveframe_category natural_source loop_ _Mol_label _Organism_name_common _NCBI_taxonomy_ID _Superkingdom _Kingdom _Genus _Species _Plasmid $Bactrx 'Bacillus acidocaldarius' 1388 Eubacteria . Alicyclobacillus acidocaldarius pTrc99A-pUC18Smal/BAP 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 $Bactrx 'recombinant technology' 'E. coli' Escherichia coli . . 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 $Bactrx 1.2 mM 1.1 1.3 . 'phosphate buffer' 50 mM . . . stop_ save_ ############################ # Computer software used # ############################ save_FELIX _Saveframe_category software _Name FELIX _Version 97 loop_ _Task 'peaks assignments' stop_ _Details . save_ ######################### # Experimental detail # ######################### ################################## # NMR Spectrometer definitions # ################################## save_NMR_spectrometer _Saveframe_category NMR_spectrometer _Manufacturer Varian _Model Unity _Field_strength 600 _Details . save_ ############################# # NMR applied experiments # ############################# save_DQF-COSY_1 _Saveframe_category NMR_applied_experiment _Experiment_name DQF-COSY _Sample_label . save_ save_TOCSY_2 _Saveframe_category NMR_applied_experiment _Experiment_name TOCSY _Sample_label . save_ save_NOESY_3 _Saveframe_category NMR_applied_experiment _Experiment_name NOESY _Sample_label . save_ save_NOESY-JR_4 _Saveframe_category NMR_applied_experiment _Experiment_name NOESY-JR _Sample_label . save_ save_DQ-2D_5 _Saveframe_category NMR_applied_experiment _Experiment_name DQ-2D _Sample_label . save_ save_MQF-COSY_6 _Saveframe_category NMR_applied_experiment _Experiment_name MQF-COSY _Sample_label . save_ save_WG-COSY_7 _Saveframe_category NMR_applied_experiment _Experiment_name WG-COSY _Sample_label . save_ save_WG-TOCSY_8 _Saveframe_category NMR_applied_experiment _Experiment_name WG-TOCSY _Sample_label . save_ save_WG-NOESY_9 _Saveframe_category NMR_applied_experiment _Experiment_name WG-NOESY _Sample_label . save_ save_3D-TOCSY-NOESY_10 _Saveframe_category NMR_applied_experiment _Experiment_name 3D-TOCSY-NOESY _Sample_label . save_ save_NMR_spec_expt__0_1 _Saveframe_category NMR_applied_experiment _Experiment_name DQF-COSY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_2 _Saveframe_category NMR_applied_experiment _Experiment_name TOCSY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_3 _Saveframe_category NMR_applied_experiment _Experiment_name NOESY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_4 _Saveframe_category NMR_applied_experiment _Experiment_name NOESY-JR _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_5 _Saveframe_category NMR_applied_experiment _Experiment_name DQ-2D _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_6 _Saveframe_category NMR_applied_experiment _Experiment_name MQF-COSY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_7 _Saveframe_category NMR_applied_experiment _Experiment_name WG-COSY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_8 _Saveframe_category NMR_applied_experiment _Experiment_name WG-TOCSY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_9 _Saveframe_category NMR_applied_experiment _Experiment_name WG-NOESY _BMRB_pulse_sequence_accession_number . _Details . save_ save_NMR_spec_expt__0_10 _Saveframe_category NMR_applied_experiment _Experiment_name 3D-TOCSY-NOESY _BMRB_pulse_sequence_accession_number . _Details . save_ ####################### # Sample conditions # ####################### save_cond_1 _Saveframe_category sample_conditions _Details . loop_ _Variable_type _Variable_value _Variable_value_error _Variable_value_units 'ionic strength' 50 . mM pH 5.8 0.2 n/a temperature 300 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 TSP H 1 'methyl protons' ppm 0.00 internal direct . internal . 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_shift_set_1 _Saveframe_category assigned_chemical_shifts _Details . loop_ _Experiment_label DQF-COSY TOCSY NOESY NOESY-JR DQ-2D MQF-COSY WG-COSY WG-TOCSY WG-NOESY 3D-TOCSY-NOESY stop_ _Sample_conditions_label $cond_1 _Chem_shift_reference_set_label $chemical_shift_reference _Mol_system_component_name thioredoxin _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 ALA HA H 4.25 . . 2 . 1 ALA HB H 1.58 . . 3 . 2 THR H H 8.72 . . 4 . 2 THR HA H 4.59 . . 5 . 2 THR HB H 3.90 . . 6 . 2 THR HG2 H 1.00 . . 7 . 3 MET H H 8.67 . . 8 . 3 MET HA H 4.67 . . 9 . 3 MET HB2 H 1.97 . . 10 . 3 MET HB3 H 1.97 . . 11 . 3 MET HG2 H 2.51 . . 12 . 3 MET HG3 H 2.37 . . 13 . 3 MET HE H 2.03 . . 14 . 4 THR H H 8.90 . . 15 . 4 THR HA H 4.60 . . 16 . 4 THR HB H 4.11 . . 17 . 4 THR HG2 H 1.31 . . 18 . 5 LEU H H 9.03 . . 19 . 5 LEU HA H 5.22 . . 20 . 5 LEU HB2 H 1.68 . . 21 . 5 LEU HB3 H 1.53 . . 22 . 5 LEU HG H 1.48 . . 23 . 5 LEU HD1 H 0.68 . . 24 . 5 LEU HD2 H 0.47 . . 25 . 6 THR H H 8.97 . . 26 . 6 THR HA H 4.71 . . 27 . 6 THR HB H 4.40 . . 28 . 6 THR HG2 H 1.21 . . 29 . 7 ASP H H 8.78 . . 30 . 7 ASP HA H 3.84 . . 31 . 7 ASP HB2 H 2.79 . . 32 . 7 ASP HB3 H 2.62 . . 33 . 8 ALA H H 8.01 . . 34 . 8 ALA HA H 4.35 . . 35 . 8 ALA HB H 1.43 . . 36 . 9 ASN H H 7.64 . . 37 . 9 ASN HA H 5.08 . . 38 . 9 ASN HB2 H 3.30 . . 39 . 9 ASN HB3 H 2.61 . . 40 . 9 ASN HD21 H 6.31 . . 41 . 9 ASN HD22 H 7.33 . . 42 . 10 PHE H H 8.14 . . 43 . 10 PHE HA H 3.77 . . 44 . 10 PHE HB2 H 3.02 . . 45 . 10 PHE HB3 H 3.39 . . 46 . 10 PHE HD1 H 6.90 . . 47 . 10 PHE HD2 H 6.90 . . 48 . 10 PHE HE1 H 6.78 . . 49 . 10 PHE HE2 H 6.78 . . 50 . 10 PHE HZ H 6.56 . . 51 . 11 GLN H H 8.56 . . 52 . 11 GLN HA H 3.73 . . 53 . 11 GLN HB2 H 2.10 . . 54 . 11 GLN HB3 H 2.10 . . 55 . 11 GLN HG2 H 2.58 . . 56 . 11 GLN HG3 H 2.58 . . 57 . 11 GLN HE21 H 6.94 . . 58 . 11 GLN HE22 H 7.76 . . 59 . 12 GLN H H 7.87 . . 60 . 12 GLN HA H 3.98 . . 61 . 12 GLN HB2 H 2.07 . . 62 . 12 GLN HB3 H 2.07 . . 63 . 12 GLN HG2 H 2.39 . . 64 . 12 GLN HG3 H 2.39 . . 65 . 13 ALA H H 7.60 . . 66 . 13 ALA HA H 4.00 . . 67 . 13 ALA HB H 1.22 . . 68 . 14 ILE H H 6.94 . . 69 . 14 ILE HA H 4.00 . . 70 . 14 ILE HB H 1.49 . . 71 . 14 ILE HG12 H 0.87 . . 72 . 14 ILE HG13 H 0.87 . . 73 . 14 ILE HD1 H 0.37 . . 74 . 15 GLN H H 6.95 . . 75 . 15 GLN HA H 4.20 . . 76 . 15 GLN HB2 H 1.93 . . 77 . 15 GLN HB3 H 1.93 . . 78 . 15 GLN HG2 H 2.51 . . 79 . 15 GLN HG3 H 2.51 . . 80 . 16 GLY H H 7.77 . . 81 . 16 GLY HA2 H 4.08 . . 82 . 16 GLY HA3 H 4.08 . . 83 . 17 ASP H H 8.27 . . 84 . 17 ASP HA H 4.48 . . 85 . 17 ASP HB2 H 2.70 . . 86 . 17 ASP HB3 H 2.64 . . 87 . 18 LYS H H 7.68 . . 88 . 18 LYS HA H 4.77 . . 89 . 18 LYS HB2 H 1.78 . . 90 . 18 LYS HB3 H 1.68 . . 91 . 18 LYS HG2 H 1.52 . . 92 . 18 LYS HG3 H 1.52 . . 93 . 18 LYS HD2 H 1.39 . . 94 . 18 LYS HD3 H 1.39 . . 95 . 18 LYS HE2 H 2.80 . . 96 . 18 LYS HE3 H 2.80 . . 97 . 19 PRO HA H 4.98 . . 98 . 19 PRO HB2 H 2.32 . . 99 . 19 PRO HB3 H 1.94 . . 100 . 19 PRO HG2 H 2.01 . . 101 . 19 PRO HG3 H 1.92 . . 102 . 19 PRO HD2 H 4.13 . . 103 . 19 PRO HD3 H 4.07 . . 104 . 20 VAL H H 8.48 . . 105 . 20 VAL HA H 4.98 . . 106 . 20 VAL HB H 1.82 . . 107 . 20 VAL HG1 H 0.89 . . 108 . 20 VAL HG2 H 0.57 . . 109 . 21 LEU H H 9.38 . . 110 . 21 LEU HA H 5.07 . . 111 . 21 LEU HB2 H 2.08 . . 112 . 21 LEU HB3 H 2.08 . . 113 . 21 LEU HG H 0.71 . . 114 . 21 LEU HD1 H 1.11 . . 115 . 21 LEU HD2 H 0.57 . . 116 . 22 VAL H H 9.43 . . 117 . 22 VAL HA H 4.43 . . 118 . 22 VAL HB H 2.24 . . 119 . 22 VAL HG1 H 0.81 . . 120 . 22 VAL HG2 H 0.20 . . 121 . 23 ASP H H 8.92 . . 122 . 23 ASP HA H 5.08 . . 123 . 23 ASP HB2 H 2.94 . . 124 . 23 ASP HB3 H 2.32 . . 125 . 24 PHE H H 9.41 . . 126 . 24 PHE HA H 5.40 . . 127 . 24 PHE HB2 H 3.43 . . 128 . 24 PHE HB3 H 2.65 . . 129 . 24 PHE HD1 H 7.36 . . 130 . 24 PHE HD2 H 7.36 . . 131 . 24 PHE HE1 H 6.98 . . 132 . 24 PHE HE2 H 6.98 . . 133 . 24 PHE HZ H 6.82 . . 134 . 25 TRP H H 8.40 . . 135 . 25 TRP HA H 5.07 . . 136 . 25 TRP HB2 H 3.08 . . 137 . 25 TRP HB3 H 2.95 . . 138 . 25 TRP HD1 H 7.31 . . 139 . 25 TRP HE1 H 10.76 . . 140 . 25 TRP HE3 H 7.06 . . 141 . 25 TRP HZ2 H 7.64 . . 142 . 25 TRP HZ3 H 6.87 . . 143 . 25 TRP HH2 H 7.27 . . 144 . 26 ALA H H 6.50 . . 145 . 26 ALA HA H 3.42 . . 146 . 26 ALA HB H 0.36 . . 147 . 27 ALA H H 9.18 . . 148 . 27 ALA HA H 4.20 . . 149 . 27 ALA HB H 1.53 . . 150 . 28 TRP H H 6.72 . . 151 . 28 TRP HA H 4.58 . . 152 . 28 TRP HB2 H 3.68 . . 153 . 28 TRP HB3 H 3.19 . . 154 . 28 TRP HD1 H 7.41 . . 155 . 28 TRP HE1 H 11.67 . . 156 . 28 TRP HE3 H 7.38 . . 157 . 28 TRP HZ2 H 7.37 . . 158 . 28 TRP HZ3 H 7.12 . . 159 . 28 TRP HH2 H 7.37 . . 160 . 29 CYS H H 6.74 . . 161 . 29 CYS HA H 4.71 . . 162 . 29 CYS HB2 H 3.07 . . 163 . 29 CYS HB3 H 2.89 . . 164 . 30 GLY H H 9.61 . . 165 . 30 GLY HA2 H 4.20 . . 166 . 30 GLY HA3 H 3.98 . . 167 . 31 PRO HA H 4.30 . . 168 . 31 PRO HB2 H 2.50 . . 169 . 31 PRO HB3 H 1.75 . . 170 . 31 PRO HG2 H 2.20 . . 171 . 31 PRO HG3 H 2.16 . . 172 . 31 PRO HD2 H 3.53 . . 173 . 31 PRO HD3 H 3.53 . . 174 . 32 CYS H H 8.21 . . 175 . 32 CYS HA H 4.46 . . 176 . 32 CYS HB2 H 4.36 . . 177 . 32 CYS HB3 H 3.36 . . 178 . 33 ARG H H 8.08 . . 179 . 33 ARG HA H 4.15 . . 180 . 33 ARG HB2 H 2.19 . . 181 . 33 ARG HB3 H 2.03 . . 182 . 33 ARG HG2 H 1.71 . . 183 . 33 ARG HG3 H 1.71 . . 184 . 33 ARG HD2 H 3.34 . . 185 . 33 ARG HD3 H 3.46 . . 186 . 33 ARG HE H 7.40 . . 187 . 34 MET H H 7.73 . . 188 . 34 MET HA H 4.30 . . 189 . 34 MET HB2 H 2.10 . . 190 . 34 MET HB3 H 2.10 . . 191 . 34 MET HG2 H 2.61 . . 192 . 34 MET HG3 H 2.55 . . 193 . 34 MET HE H 1.84 . . 194 . 35 MET H H 7.68 . . 195 . 35 MET HA H 4.57 . . 196 . 35 MET HB2 H 2.22 . . 197 . 35 MET HB3 H 2.14 . . 198 . 35 MET HG2 H 2.70 . . 199 . 35 MET HG3 H 2.55 . . 200 . 35 MET HE H 2.00 . . 201 . 36 ALA H H 7.41 . . 202 . 36 ALA HA H 4.00 . . 203 . 36 ALA HB H 1.40 . . 204 . 37 PRO HA H 4.52 . . 205 . 37 PRO HB2 H 2.01 . . 206 . 37 PRO HB3 H 1.91 . . 207 . 37 PRO HG2 H 2.11 . . 208 . 37 PRO HG3 H 2.11 . . 209 . 37 PRO HD2 H 3.42 . . 210 . 37 PRO HD3 H 3.42 . . 211 . 38 VAL H H 6.61 . . 212 . 38 VAL HA H 3.82 . . 213 . 38 VAL HB H 2.35 . . 214 . 38 VAL HG1 H 1.25 . . 215 . 38 VAL HG2 H 0.94 . . 216 . 39 LEU H H 8.16 . . 217 . 39 LEU HA H 3.92 . . 218 . 39 LEU HB2 H 1.74 . . 219 . 39 LEU HB3 H 1.40 . . 220 . 39 LEU HG H 1.62 . . 221 . 39 LEU HD1 H 0.68 . . 222 . 39 LEU HD2 H 0.61 . . 223 . 40 GLU H H 7.82 . . 224 . 40 GLU HA H 3.97 . . 225 . 40 GLU HB2 H 2.14 . . 226 . 40 GLU HB3 H 2.14 . . 227 . 40 GLU HG2 H 2.32 . . 228 . 40 GLU HG3 H 2.32 . . 229 . 41 GLU H H 7.80 . . 230 . 41 GLU HA H 4.10 . . 231 . 41 GLU HB2 H 2.23 . . 232 . 41 GLU HB3 H 2.13 . . 233 . 41 GLU HG2 H 2.50 . . 234 . 41 GLU HG3 H 2.50 . . 235 . 42 PHE H H 8.57 . . 236 . 42 PHE HA H 4.14 . . 237 . 42 PHE HB2 H 3.23 . . 238 . 42 PHE HB3 H 3.11 . . 239 . 42 PHE HD1 H 7.08 . . 240 . 42 PHE HD2 H 7.08 . . 241 . 42 PHE HE1 H 6.82 . . 242 . 42 PHE HE2 H 6.82 . . 243 . 42 PHE HZ H 6.74 . . 244 . 43 ALA H H 8.74 . . 245 . 43 ALA HA H 3.93 . . 246 . 43 ALA HB H 1.52 . . 247 . 44 GLU H H 7.58 . . 248 . 44 GLU HA H 4.08 . . 249 . 44 GLU HB2 H 2.09 . . 250 . 44 GLU HB3 H 2.09 . . 251 . 44 GLU HG2 H 2.32 . . 252 . 44 GLU HG3 H 2.42 . . 253 . 45 ALA H H 7.71 . . 254 . 45 ALA HA H 4.02 . . 255 . 45 ALA HB H 0.95 . . 256 . 46 HIS H H 7.32 . . 257 . 46 HIS HA H 5.08 . . 258 . 46 HIS HB2 H 3.10 . . 259 . 46 HIS HB3 H 2.15 . . 260 . 46 HIS HD2 H 5.73 . . 261 . 46 HIS HE1 H 8.43 . . 262 . 47 ALA H H 7.15 . . 263 . 47 ALA HA H 4.46 . . 264 . 47 ALA HB H 1.57 . . 265 . 48 ASP H H 8.86 . . 266 . 48 ASP HA H 4.37 . . 267 . 48 ASP HB2 H 2.74 . . 268 . 48 ASP HB3 H 2.69 . . 269 . 49 LYS H H 7.86 . . 270 . 49 LYS HA H 4.58 . . 271 . 49 LYS HB2 H 1.73 . . 272 . 49 LYS HB3 H 1.53 . . 273 . 49 LYS HG2 H 1.40 . . 274 . 49 LYS HG3 H 1.40 . . 275 . 49 LYS HD2 H 1.68 . . 276 . 49 LYS HD3 H 1.68 . . 277 . 49 LYS HE2 H 2.94 . . 278 . 49 LYS HE3 H 2.94 . . 279 . 50 VAL H H 7.95 . . 280 . 50 VAL HA H 4.71 . . 281 . 50 VAL HB H 1.95 . . 282 . 50 VAL HG1 H 0.86 . . 283 . 50 VAL HG2 H 0.82 . . 284 . 51 THR H H 8.72 . . 285 . 51 THR HA H 4.48 . . 286 . 51 THR HB H 4.02 . . 287 . 51 THR HG2 H 1.23 . . 288 . 52 VAL H H 9.11 . . 289 . 52 VAL HA H 4.92 . . 290 . 52 VAL HB H 2.04 . . 291 . 52 VAL HG1 H 1.11 . . 292 . 52 VAL HG2 H 0.95 . . 293 . 53 ALA H H 9.43 . . 294 . 53 ALA HA H 5.45 . . 295 . 53 ALA HB H 1.13 . . 296 . 54 LYS H H 8.65 . . 297 . 54 LYS HA H 5.13 . . 298 . 54 LYS HB2 H 1.82 . . 299 . 54 LYS HB3 H 1.82 . . 300 . 54 LYS HG2 H 1.39 . . 301 . 54 LYS HG3 H 1.24 . . 302 . 54 LYS HD2 H 1.21 . . 303 . 54 LYS HD3 H 1.21 . . 304 . 54 LYS HE2 H 2.30 . . 305 . 54 LYS HE3 H 2.30 . . 306 . 55 LEU H H 8.92 . . 307 . 55 LEU HA H 4.60 . . 308 . 55 LEU HB2 H 1.20 . . 309 . 55 LEU HB3 H 0.80 . . 310 . 55 LEU HG H 1.36 . . 311 . 55 LEU HD1 H 0.76 . . 312 . 55 LEU HD2 H 0.66 . . 313 . 56 ASN H H 9.01 . . 314 . 56 ASN HA H 3.93 . . 315 . 56 ASN HB2 H 2.59 . . 316 . 56 ASN HB3 H 2.31 . . 317 . 56 ASN HD21 H 7.00 . . 318 . 56 ASN HD22 H 5.60 . . 319 . 57 VAL H H 8.76 . . 320 . 57 VAL HA H 4.60 . . 321 . 57 VAL HB H 2.24 . . 322 . 57 VAL HG1 H 1.12 . . 323 . 57 VAL HG2 H 0.60 . . 324 . 58 ASP H H 7.50 . . 325 . 58 ASP HA H 4.50 . . 326 . 58 ASP HB2 H 2.86 . . 327 . 58 ASP HB3 H 2.80 . . 328 . 59 GLU H H 7.18 . . 329 . 59 GLU HA H 4.20 . . 330 . 59 GLU HB2 H 1.72 . . 331 . 59 GLU HB3 H 1.72 . . 332 . 59 GLU HG2 H 2.24 . . 333 . 59 GLU HG3 H 2.11 . . 334 . 60 ASN H H 6.93 . . 335 . 60 ASN HA H 5.43 . . 336 . 60 ASN HB2 H 2.87 . . 337 . 60 ASN HB3 H 2.61 . . 338 . 60 ASN HD21 H 8.32 . . 339 . 60 ASN HD22 H 7.32 . . 340 . 61 PRO HA H 4.70 . . 341 . 61 PRO HB2 H 2.10 . . 342 . 61 PRO HB3 H 2.10 . . 343 . 61 PRO HG2 H 2.46 . . 344 . 61 PRO HG3 H 2.14 . . 345 . 61 PRO HD2 H 3.93 . . 346 . 61 PRO HD3 H 3.40 . . 347 . 62 GLU H H 9.44 . . 348 . 62 GLU HA H 4.12 . . 349 . 62 GLU HB2 H 1.99 . . 350 . 62 GLU HB3 H 1.19 . . 351 . 62 GLU HG2 H 2.35 . . 352 . 62 GLU HG3 H 2.24 . . 353 . 63 THR H H 9.66 . . 354 . 63 THR HA H 3.70 . . 355 . 63 THR HB H 3.73 . . 356 . 63 THR HG2 H 0.37 . . 357 . 64 THR H H 7.12 . . 358 . 64 THR HA H 3.60 . . 359 . 64 THR HB H 4.10 . . 360 . 64 THR HG2 H 0.77 . . 361 . 65 SER H H 7.74 . . 362 . 65 SER HA H 4.37 . . 363 . 65 SER HB2 H 4.03 . . 364 . 65 SER HB3 H 4.03 . . 365 . 66 GLN H H 8.16 . . 366 . 66 GLN HA H 3.86 . . 367 . 66 GLN HB2 H 1.74 . . 368 . 66 GLN HB3 H 1.74 . . 369 . 66 GLN HG2 H 2.04 . . 370 . 66 GLN HG3 H 2.04 . . 371 . 67 PHE H H 7.23 . . 372 . 67 PHE HA H 4.50 . . 373 . 67 PHE HB2 H 3.31 . . 374 . 67 PHE HB3 H 2.30 . . 375 . 67 PHE HD1 H 7.51 . . 376 . 67 PHE HD2 H 7.51 . . 377 . 67 PHE HE1 H 7.41 . . 378 . 67 PHE HE2 H 7.41 . . 379 . 67 PHE HZ H 7.67 . . 380 . 68 GLY H H 7.67 . . 381 . 68 GLY HA2 H 3.92 . . 382 . 68 GLY HA3 H 3.78 . . 383 . 69 ILE H H 7.34 . . 384 . 69 ILE HA H 3.78 . . 385 . 69 ILE HB H 1.91 . . 386 . 69 ILE HG12 H 1.27 . . 387 . 69 ILE HG13 H 1.27 . . 388 . 69 ILE HG2 H 0.65 . . 389 . 70 MET H H 8.57 . . 390 . 70 MET HA H 4.70 . . 391 . 70 MET HB2 H 2.20 . . 392 . 70 MET HB3 H 2.20 . . 393 . 70 MET HG2 H 2.69 . . 394 . 70 MET HG3 H 2.69 . . 395 . 71 SER H H 7.84 . . 396 . 71 SER HA H 4.83 . . 397 . 71 SER HB2 H 3.73 . . 398 . 71 SER HB3 H 3.73 . . 399 . 72 ILE H H 8.41 . . 400 . 72 ILE HA H 4.73 . . 401 . 72 ILE HB H 1.92 . . 402 . 72 ILE HG12 H 1.49 . . 403 . 72 ILE HG13 H 1.20 . . 404 . 72 ILE HG2 H 0.76 . . 405 . 72 ILE HD1 H 0.35 . . 406 . 73 PRO HA H 5.16 . . 407 . 73 PRO HB2 H 2.73 . . 408 . 73 PRO HB3 H 2.00 . . 409 . 73 PRO HG2 H 1.77 . . 410 . 73 PRO HG3 H 1.50 . . 411 . 73 PRO HD2 H 3.69 . . 412 . 73 PRO HD3 H 3.53 . . 413 . 74 THR H H 8.05 . . 414 . 74 THR HA H 4.97 . . 415 . 74 THR HB H 3.93 . . 416 . 74 THR HG2 H 1.11 . . 417 . 75 LEU H H 9.41 . . 418 . 75 LEU HA H 6.02 . . 419 . 75 LEU HB2 H 1.72 . . 420 . 75 LEU HB3 H 1.63 . . 421 . 75 LEU HG H 1.79 . . 422 . 75 LEU HD1 H 1.27 . . 423 . 75 LEU HD2 H 0.77 . . 424 . 76 ILE H H 8.97 . . 425 . 76 ILE HA H 4.86 . . 426 . 76 ILE HB H 1.72 . . 427 . 76 ILE HG12 H 1.64 . . 428 . 76 ILE HG13 H 1.70 . . 429 . 76 ILE HG2 H 0.08 . . 430 . 76 ILE HD1 H 0.61 . . 431 . 77 LEU H H 8.59 . . 432 . 77 LEU HA H 5.12 . . 433 . 77 LEU HB2 H 1.87 . . 434 . 77 LEU HB3 H 1.00 . . 435 . 77 LEU HG H 1.42 . . 436 . 77 LEU HD1 H 0.87 . . 437 . 77 LEU HD2 H 0.87 . . 438 . 78 PHE H H 9.91 . . 439 . 78 PHE HA H 5.23 . . 440 . 78 PHE HB2 H 3.00 . . 441 . 78 PHE HB3 H 2.80 . . 442 . 78 PHE HD1 H 7.15 . . 443 . 78 PHE HD2 H 7.15 . . 444 . 78 PHE HE1 H 6.94 . . 445 . 78 PHE HE2 H 6.94 . . 446 . 78 PHE HZ H 6.22 . . 447 . 79 LYS H H 8.68 . . 448 . 79 LYS HA H 4.71 . . 449 . 79 LYS HB2 H 1.81 . . 450 . 79 LYS HB3 H 1.69 . . 451 . 79 LYS HG2 H 1.43 . . 452 . 79 LYS HG3 H 1.38 . . 453 . 79 LYS HD2 H 1.57 . . 454 . 79 LYS HD3 H 1.57 . . 455 . 79 LYS HE2 H 3.03 . . 456 . 79 LYS HE3 H 3.03 . . 457 . 80 GLY H H 9.31 . . 458 . 80 GLY HA2 H 4.06 . . 459 . 80 GLY HA3 H 3.91 . . 460 . 81 GLY H H 8.90 . . 461 . 81 GLY HA2 H 4.33 . . 462 . 81 GLY HA3 H 3.53 . . 463 . 82 ARG H H 7.72 . . 464 . 82 ARG HA H 5.25 . . 465 . 82 ARG HB2 H 1.95 . . 466 . 82 ARG HB3 H 1.87 . . 467 . 82 ARG HG2 H 1.73 . . 468 . 82 ARG HG3 H 1.73 . . 469 . 82 ARG HD2 H 3.30 . . 470 . 82 ARG HD3 H 3.30 . . 471 . 82 ARG HE H 7.23 . . 472 . 83 PRO HA H 4.00 . . 473 . 83 PRO HB2 H 2.42 . . 474 . 83 PRO HB3 H 2.09 . . 475 . 83 PRO HG2 H 1.52 . . 476 . 83 PRO HG3 H 1.52 . . 477 . 83 PRO HD2 H 4.10 . . 478 . 83 PRO HD3 H 3.92 . . 479 . 84 VAL H H 9.21 . . 480 . 84 VAL HA H 4.43 . . 481 . 84 VAL HB H 2.18 . . 482 . 84 VAL HG1 H 1.00 . . 483 . 84 VAL HG2 H 0.87 . . 484 . 85 LYS H H 7.27 . . 485 . 85 LYS HA H 4.53 . . 486 . 85 LYS HB2 H 1.77 . . 487 . 85 LYS HB3 H 1.60 . . 488 . 85 LYS HG2 H 1.35 . . 489 . 85 LYS HG3 H 1.35 . . 490 . 85 LYS HD2 H 1.64 . . 491 . 85 LYS HD3 H 1.64 . . 492 . 85 LYS HE2 H 3.49 . . 493 . 85 LYS HE3 H 3.03 . . 494 . 86 GLN H H 8.62 . . 495 . 86 GLN HA H 5.30 . . 496 . 86 GLN HB2 H 1.92 . . 497 . 86 GLN HB3 H 1.88 . . 498 . 86 GLN HG2 H 2.15 . . 499 . 86 GLN HG3 H 2.07 . . 500 . 87 LEU H H 9.42 . . 501 . 87 LEU HA H 4.79 . . 502 . 87 LEU HB2 H 1.42 . . 503 . 87 LEU HB3 H 1.39 . . 504 . 87 LEU HG H 1.44 . . 505 . 87 LEU HD1 H 0.72 . . 506 . 87 LEU HD2 H 0.59 . . 507 . 88 ILE H H 8.73 . . 508 . 88 ILE HA H 4.60 . . 509 . 88 ILE HB H 1.80 . . 510 . 88 ILE HG12 H 0.90 . . 511 . 88 ILE HG13 H 0.90 . . 512 . 88 ILE HG2 H 0.86 . . 513 . 88 ILE HD1 H 0.77 . . 514 . 89 GLY H H 8.15 . . 515 . 89 GLY HA2 H 4.36 . . 516 . 89 GLY HA3 H 3.69 . . 517 . 90 TYR H H 8.50 . . 518 . 90 TYR HA H 4.11 . . 519 . 90 TYR HB2 H 2.95 . . 520 . 90 TYR HB3 H 2.84 . . 521 . 90 TYR HD1 H 6.94 . . 522 . 90 TYR HD2 H 6.94 . . 523 . 90 TYR HE1 H 6.62 . . 524 . 90 TYR HE2 H 6.62 . . 525 . 91 GLN H H 6.82 . . 526 . 91 GLN HA H 4.52 . . 527 . 91 GLN HB2 H 1.74 . . 528 . 91 GLN HB3 H 1.40 . . 529 . 91 GLN HG2 H 2.43 . . 530 . 91 GLN HG3 H 2.43 . . 531 . 91 GLN HE21 H 6.36 . . 532 . 91 GLN HE22 H 6.25 . . 533 . 92 PRO HA H 4.36 . . 534 . 92 PRO HB2 H 2.53 . . 535 . 92 PRO HB3 H 2.14 . . 536 . 92 PRO HG2 H 2.10 . . 537 . 92 PRO HG3 H 1.90 . . 538 . 92 PRO HD2 H 3.45 . . 539 . 92 PRO HD3 H 3.40 . . 540 . 93 LYS H H 8.93 . . 541 . 93 LYS HA H 3.60 . . 542 . 93 LYS HB2 H 2.04 . . 543 . 93 LYS HB3 H 1.81 . . 544 . 93 LYS HG2 H 0.93 . . 545 . 93 LYS HG3 H 0.93 . . 546 . 93 LYS HD2 H 1.49 . . 547 . 93 LYS HD3 H 1.49 . . 548 . 93 LYS HE2 H 2.98 . . 549 . 93 LYS HE3 H 2.98 . . 550 . 94 GLU H H 9.52 . . 551 . 94 GLU HA H 4.18 . . 552 . 94 GLU HB2 H 2.07 . . 553 . 94 GLU HB3 H 2.07 . . 554 . 94 GLU HG2 H 2.46 . . 555 . 94 GLU HG3 H 2.37 . . 556 . 95 GLN H H 7.20 . . 557 . 95 GLN HA H 4.33 . . 558 . 95 GLN HB2 H 2.20 . . 559 . 95 GLN HB3 H 2.01 . . 560 . 95 GLN HG2 H 2.47 . . 561 . 95 GLN HG3 H 2.47 . . 562 . 96 LEU H H 7.95 . . 563 . 96 LEU HA H 3.63 . . 564 . 96 LEU HB2 H 1.39 . . 565 . 96 LEU HB3 H 1.01 . . 566 . 96 LEU HG H 1.06 . . 567 . 96 LEU HD1 H 0.25 . . 568 . 96 LEU HD2 H 0.13 . . 569 . 97 GLU H H 8.15 . . 570 . 97 GLU HA H 3.76 . . 571 . 97 GLU HB2 H 2.21 . . 572 . 97 GLU HB3 H 2.11 . . 573 . 97 GLU HG2 H 2.62 . . 574 . 97 GLU HG3 H 2.62 . . 575 . 98 ALA H H 7.50 . . 576 . 98 ALA HA H 4.19 . . 577 . 98 ALA HB H 1.54 . . 578 . 99 GLN H H 8.33 . . 579 . 99 GLN HA H 4.18 . . 580 . 99 GLN HB2 H 2.10 . . 581 . 99 GLN HB3 H 1.90 . . 582 . 99 GLN HG2 H 2.49 . . 583 . 99 GLN HG3 H 2.49 . . 584 . 100 LEU H H 7.49 . . 585 . 100 LEU HA H 4.60 . . 586 . 100 LEU HB2 H 1.74 . . 587 . 100 LEU HB3 H 1.71 . . 588 . 100 LEU HG H 1.63 . . 589 . 100 LEU HD1 H 0.74 . . 590 . 100 LEU HD2 H 0.63 . . 591 . 101 ALA H H 7.15 . . 592 . 101 ALA HA H 3.90 . . 593 . 101 ALA HB H 1.52 . . 594 . 102 ASP H H 8.61 . . 595 . 102 ASP HA H 4.37 . . 596 . 102 ASP HB2 H 2.68 . . 597 . 102 ASP HB3 H 2.64 . . 598 . 103 VAL H H 7.57 . . 599 . 103 VAL HA H 4.42 . . 600 . 103 VAL HB H 2.32 . . 601 . 103 VAL HG1 H 0.93 . . 602 . 103 VAL HG2 H 0.93 . . 603 . 104 LEU H H 7.39 . . 604 . 104 LEU HA H 4.30 . . 605 . 104 LEU HB2 H 1.64 . . 606 . 104 LEU HB3 H 1.64 . . 607 . 104 LEU HG H 1.76 . . 608 . 104 LEU HD1 H 0.48 . . 609 . 104 LEU HD2 H 0.50 . . 610 . 105 GLN H H 7.30 . . 611 . 105 GLN HA H 4.08 . . 612 . 105 GLN HB2 H 2.10 . . 613 . 105 GLN HB3 H 2.01 . . 614 . 105 GLN HG2 H 2.37 . . 615 . 105 GLN HG3 H 2.37 . . stop_ save_