data_5028 ####################### # Entry information # ####################### save_entry_information _Saveframe_category entry_information _Entry_title ; Solution Structure of MCOTI-II, a Macrocyclic Trypsin Inhibitor ; _BMRB_accession_number 5028 _BMRB_flat_file_name bmr5028.str _Entry_type original _Submission_date 2001-05-22 _Accession_date 2001-05-22 _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 Felizmenio-Quimio M. F. . 2 Daly N. L. . 3 Craik D. J. . stop_ loop_ _Saveframe_category_type _Saveframe_category_type_count assigned_chemical_shifts 1 stop_ loop_ _Data_type _Data_type_count "1H chemical shifts" 186 stop_ loop_ _Revision_date _Revision_keyword _Revision_author _Revision_detail 2001-05-23 original BMRB . stop_ _Original_release_date 2001-05-22 save_ ############################# # Citation for this entry # ############################# save_entry_citation _Saveframe_category entry_citation _Citation_full . _Citation_title ; Circular Proteins in Plants: Solution Structure of a Novel Macrocyclic Trypsin Inhibitor from Momordica cochinchinensis ; _Citation_status 'in press' _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 Felizmenio-Quimio M. F. . 2 Daly N. L. . 3 Craik D. J. . stop_ _Journal_abbreviation 'J. Biol. Chem.' _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 2001 _Details . loop_ _Keyword 'Cyclic cystine knot' beta-hairpin 'circular protein' cyclotide stop_ save_ ####################################### # Cited references within the entry # ####################################### save_reference_1 _Saveframe_category citation _Citation_full ; Hernandez JF, Gagnon J, Chiche L, Nguyen TM, Andrieu JP, Heitz A, Trinh Hong T, Pham TT, Le Nguyen D. Squash trypsin inhibitors from Momordica cochinchinensis exhibit an atypical macrocyclic structure. Biochemistry. 2000 May 16;39(19):5722-30. ; _Citation_title ; Squash trypsin inhibitors from Momordica cochinchinensis exhibit an atypical macrocyclic structure. ; _Citation_status published _Citation_type journal _CAS_abstract_code . _MEDLINE_UI_code . _PubMed_ID 10801322 loop_ _Author_ordinal _Author_family_name _Author_given_name _Author_middle_initials _Author_family_title 1 Hernandez 'J. F.' F. . 2 Gagnon J. . . 3 Chiche L. . . 4 Nguyen 'T. M.' M. . 5 Andrieu 'J. P.' P. . 6 Heitz A. . . 7 'Trinh Hong' T. . . 8 Pham 'T. T.' T. . 9 'Le Nguyen' D. . . stop_ _Journal_abbreviation Biochemistry _Journal_name_full Biochemistry _Journal_volume 39 _Journal_issue 19 _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 5722 _Page_last 5730 _Year 2000 _Details ; Three trypsin inhibitors (TIs), from the seeds of the squash Momordica cochinchinensis (MCo), have been isolated and purified using gel filtration, ion exchange chromatography, and reverse-phase HPLC. Their sequences could be determined only after proteolytic cleavages. In the case of MCoTI-I and -II, it was shown that their polypeptide backbones are cyclic, a structure that has never been described in squash TIs. They contain 34 amino acid residues with 3 disulfide bridges and measured molecular masses of 3453.0 and 3480.7, respectively. They are the largest known macrocyclic peptides containing disulfide bridges. Their sequences show strong homology to other squash TIs, suggesting a similar three-dimensional structure and an analogous mechanism of action. A model of MCoTI-II was constructed by analogy to the crystal structure of the complex between bovine trypsin and CMTI-I, indicating that the linker connecting the two termini is flexible and does not impose significant geometrical constraints. This flexibility allows an Asp-Gly peptide bond rearrangement to occur in this region, giving rise to two isoforms of MCoTI-II. Although the importance of cyclization is not clear, it might confer increased stability and resistance to proteolysis. A minor species, MCoTI-III, was also characterized as containing 30 amino acid residues with a molecular mass of 3379.6. This component possesses a linear backbone with a blocked N-terminus. MCoTIs represent interesting candidates for drug design, either by changing their specificity of inhibition or by using their structure as natural scaffolds bearing new binding activities. ; save_ ################################## # Molecular system description # ################################## save_system_MCoTI_II _Saveframe_category molecular_system _Mol_system_name 'TRYPSIN INHIBITOR II' _Abbreviation_common 'MCoTI II' _Enzyme_commission_number . loop_ _Mol_system_component_name _Mol_label 'MCoTI II' $MCoTI_II stop_ _System_molecular_weight . _System_physical_state native _System_oligomer_state monomer _System_paramagnetic no _System_thiol_state 'all disulfide bound' loop_ _Biological_function 'trypsin inhibitor' stop_ _Database_query_date . _Details . save_ ######################## # Monomeric polymers # ######################## save_MCoTI_II _Saveframe_category monomeric_polymer _Mol_type polymer _Mol_polymer_class protein _Name_common 'MCoTI II' _Abbreviation_common 'MCoTI II' _Molecular_mass 3453 _Mol_thiol_state 'all disulfide bound' _Details . ############################## # Polymer residue sequence # ############################## _Residue_count 34 _Mol_residue_sequence ; SGSDGGVCPKILKKCRRDSD CPGACICRGNGYCG ; loop_ _Residue_seq_code _Residue_label 1 SER 2 GLY 3 SER 4 ASP 5 GLY 6 GLY 7 VAL 8 CYS 9 PRO 10 LYS 11 ILE 12 LEU 13 LYS 14 LYS 15 CYS 16 ARG 17 ARG 18 ASP 19 SER 20 ASP 21 CYS 22 PRO 23 GLY 24 ALA 25 CYS 26 ILE 27 CYS 28 ARG 29 GLY 30 ASN 31 GLY 32 TYR 33 CYS 34 GLY stop_ _Sequence_homology_query_date 2008-08-19 _Sequence_homology_query_revised_last_date 2008-08-19 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 BMRB 5176 MCoTI-II 100.00 34 100.00 100.00 4.33e-10 BMRB 7361 lin_MCoTI 82.35 29 100.00 100.00 3.98e-07 PDB 1HA9 'Solution Structure Of The Squash Trypsin Inhibitor Mcoti-Ii, Nmr, 30 Structures.' 97.06 34 100.00 100.00 1.23e-09 PDB 1IB9 'Solution Structure Of Mcoti-Ii, A Macrocyclic Trypsin Inhibitor' 97.06 34 100.00 100.00 1.23e-09 PDB 2IT8 'Solution Structure Of A Linear Analog Of The Cyclic Squash Trypsin Inhibitor Mcoti-Ii' 85.29 30 100.00 100.00 1.29e-07 PDB 2PO8 'The Structure Of A Two-Disulfide Intermediate Of Mcoti-Ii' 79.41 34 100.00 100.00 1.07e-06 SWISS-PROT P82409 'Trypsin inhibitor 2 (Trypsin inhibitor II) (MCoTI-II)' 100.00 34 100.00 100.00 4.33e-10 stop_ save_ #################### # Natural source # #################### save_natural_source _Saveframe_category natural_source loop_ _Mol_label _Organism_name_common _NCBI_taxonomy_ID _Superkingdom _Kingdom _Genus _Species _Tissue $MCoTI_II 'spiny bitter cucumber' 3674 Eukaryota Viridiplantae Momordica cochinchinensis seed 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 $MCoTI_II 'purified from the natural source' . . . . . 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 $MCoTI_II 1 mM . H2O 90 % . D2O 10 % . stop_ save_ ############################ # Computer software used # ############################ save_XWINNMR _Saveframe_category software _Name xwinnmr _Version 2.6 loop_ _Task collection stop_ _Details Bruker save_ save_XEASY _Saveframe_category software _Name XEASY _Version 1.3.7 loop_ _Task 'data analysis' stop_ _Details 'Eccles, Guntert, Billeter, and Wuthrich.' save_ save_DYANA _Saveframe_category software _Name DYANA _Version 1.5 loop_ _Task 'structure solution' stop_ _Details 'Guntert, Mumenthaler, and Wuthrich.' save_ save_X-PLOR _Saveframe_category software _Name X-PLOR _Version 3.851 loop_ _Task 'structure solution' stop_ _Details Brunger save_ ######################### # Experimental detail # ######################### ################################## # NMR Spectrometer definitions # ################################## save_NMR_spectrometer _Saveframe_category NMR_spectrometer _Manufacturer Bruker _Model AVANCE _Field_strength 750 _Details . save_ ############################# # NMR applied experiments # ############################# save_2D_NOESY_1 _Saveframe_category NMR_applied_experiment _Experiment_name '2D NOESY' _Sample_label $sample_1 save_ save_DQF-COSY_2 _Saveframe_category NMR_applied_experiment _Experiment_name DQF-COSY _Sample_label $sample_1 save_ save_E-COSY_3 _Saveframe_category NMR_applied_experiment _Experiment_name E-COSY _Sample_label $sample_1 save_ ####################### # Sample conditions # ####################### save_sample_cond_1 _Saveframe_category sample_conditions _Details . loop_ _Variable_type _Variable_value _Variable_value_error _Variable_value_units pH 3.5 0.2 n/a pressure 1 . atm temperature 293 1 K stop_ save_ #################### # NMR parameters # #################### ############################## # Assigned chemical shifts # ############################## ################################ # Chemical shift referencing # ################################ save_chemical_shift_ref_1 _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_chemical_shift_set_1 _Saveframe_category assigned_chemical_shifts _Details . loop_ _Experiment_label '2D NOESY' DQF-COSY E-COSY stop_ loop_ _Sample_label $sample_1 stop_ _Sample_conditions_label $sample_cond_1 _Chem_shift_reference_set_label $chemical_shift_ref_1 _Mol_system_component_name 'MCoTI II' _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 SER H H 8.852 0.02 1 2 . 1 SER HA H 4.529 0.02 1 3 . 1 SER HB2 H 3.933 0.02 1 4 . 1 SER HB3 H 3.933 0.02 1 5 . 2 GLY H H 9.189 0.02 1 6 . 2 GLY HA2 H 4.443 0.02 2 7 . 2 GLY HA3 H 3.898 0.02 2 8 . 3 SER H H 8.738 0.02 1 9 . 3 SER HA H 4.486 0.02 1 10 . 3 SER HB2 H 3.915 0.02 1 11 . 3 SER HB3 H 3.915 0.02 1 12 . 4 ASP H H 8.621 0.02 1 13 . 4 ASP HA H 4.824 0.02 1 14 . 4 ASP HB2 H 2.947 0.02 2 15 . 4 ASP HB3 H 2.862 0.02 2 16 . 5 GLY H H 8.25 0.02 1 17 . 5 GLY HA2 H 4.11 0.02 2 18 . 5 GLY HA3 H 3.834 0.02 2 19 . 6 GLY H H 8.162 0.02 1 20 . 6 GLY HA2 H 3.93 0.02 1 21 . 6 GLY HA3 H 3.93 0.02 1 22 . 7 VAL H H 8.542 0.02 1 23 . 7 VAL HA H 4.016 0.02 1 24 . 7 VAL HB H 1.991 0.02 1 25 . 7 VAL HG1 H 0.943 0.02 1 26 . 7 VAL HG2 H 0.839 0.02 1 27 . 8 CYS H H 8.75 0.02 1 28 . 8 CYS HA H 5.198 0.02 1 29 . 8 CYS HB2 H 2.875 0.02 1 30 . 8 CYS HB3 H 2.875 0.02 1 31 . 9 PRO HA H 4.371 0.02 1 32 . 9 PRO HB2 H 2.31 0.02 2 33 . 9 PRO HB3 H 1.862 0.02 2 34 . 9 PRO HG2 H 2.007 0.02 2 35 . 9 PRO HG3 H 1.947 0.02 2 36 . 9 PRO HD2 H 3.853 0.02 1 37 . 9 PRO HD3 H 3.853 0.02 1 38 . 10 LYS H H 8.29 0.02 1 39 . 10 LYS HA H 4.263 0.02 1 40 . 10 LYS HB2 H 1.829 0.02 2 41 . 10 LYS HB3 H 1.704 0.02 2 42 . 10 LYS HG2 H 1.367 0.02 1 43 . 10 LYS HG3 H 1.367 0.02 1 44 . 10 LYS HD2 H 1.481 0.02 1 45 . 10 LYS HD3 H 1.481 0.02 1 46 . 10 LYS HE2 H 2.993 0.02 1 47 . 10 LYS HE3 H 2.993 0.02 1 48 . 10 LYS HZ H 7.542 0.02 1 49 . 11 ILE H H 7.752 0.02 1 50 . 11 ILE HA H 4.387 0.02 1 51 . 11 ILE HB H 1.795 0.02 1 52 . 11 ILE HG12 H 1.356 0.02 1 53 . 11 ILE HG13 H 1.356 0.02 1 54 . 11 ILE HG2 H 0.831 0.02 1 55 . 11 ILE HD1 H 1.07 0.02 1 56 . 12 LEU H H 8.736 0.02 1 57 . 12 LEU HA H 4.548 0.02 1 58 . 12 LEU HB2 H 1.747 0.02 1 59 . 12 LEU HB3 H 1.747 0.02 1 60 . 12 LEU HG H 1.478 0.02 1 61 . 12 LEU HD1 H 0.767 0.02 1 62 . 12 LEU HD2 H 0.767 0.02 1 63 . 13 LYS H H 8.946 0.02 1 64 . 13 LYS HA H 4.592 0.02 1 65 . 13 LYS HB2 H 1.606 0.02 2 66 . 13 LYS HB3 H 1.566 0.02 2 67 . 13 LYS HG2 H 1.414 0.02 1 68 . 13 LYS HG3 H 1.414 0.02 1 69 . 13 LYS HD2 H 1.754 0.02 1 70 . 13 LYS HD3 H 1.754 0.02 1 71 . 13 LYS HE2 H 3.089 0.02 1 72 . 13 LYS HE3 H 3.089 0.02 1 73 . 13 LYS HZ H 7.626 0.02 1 74 . 14 LYS H H 8.672 0.02 1 75 . 14 LYS HA H 4.316 0.02 1 76 . 14 LYS HB2 H 1.643 0.02 2 77 . 14 LYS HB3 H 1.513 0.02 2 78 . 14 LYS HG2 H 0.937 0.02 1 79 . 14 LYS HG3 H 0.937 0.02 1 80 . 14 LYS HD2 H 1.309 0.02 1 81 . 14 LYS HD3 H 1.309 0.02 1 82 . 14 LYS HE2 H 2.93 0.02 1 83 . 14 LYS HE3 H 2.93 0.02 1 84 . 14 LYS HZ H 7.474 0.02 1 85 . 15 CYS H H 8.437 0.02 1 86 . 15 CYS HA H 4.888 0.02 1 87 . 15 CYS HB2 H 3.198 0.02 2 88 . 15 CYS HB3 H 2.932 0.02 2 89 . 16 ARG H H 9.399 0.02 1 90 . 16 ARG HA H 4.443 0.02 1 91 . 16 ARG HB2 H 1.881 0.02 2 92 . 16 ARG HB3 H 1.729 0.02 2 93 . 16 ARG HG2 H 1.63 0.02 1 94 . 16 ARG HG3 H 1.63 0.02 1 95 . 16 ARG HD2 H 3.186 0.02 1 96 . 16 ARG HD3 H 3.186 0.02 1 97 . 16 ARG HE H 7.308 0.02 1 98 . 17 ARG H H 8.081 0.02 1 99 . 17 ARG HA H 4.742 0.02 1 100 . 17 ARG HB2 H 2.034 0.02 2 101 . 17 ARG HB3 H 1.793 0.02 2 102 . 17 ARG HG2 H 1.559 0.02 2 103 . 17 ARG HG3 H 1.457 0.02 2 104 . 17 ARG HD2 H 3.153 0.02 1 105 . 17 ARG HD3 H 3.153 0.02 1 106 . 17 ARG HE H 7.169 0.02 1 107 . 18 ASP H H 9.348 0.02 1 108 . 18 ASP HA H 4.118 0.02 1 109 . 18 ASP HB2 H 3.004 0.02 2 110 . 18 ASP HB3 H 2.83 0.02 2 111 . 19 SER H H 8.217 0.02 1 112 . 19 SER HA H 4.303 0.02 1 113 . 19 SER HB2 H 4.119 0.02 1 114 . 19 SER HB3 H 3.834 0.02 1 115 . 20 ASP H H 7.739 0.02 1 116 . 20 ASP HA H 4.634 0.02 1 117 . 20 ASP HB2 H 3.039 0.02 2 118 . 20 ASP HB3 H 2.965 0.02 2 119 . 21 CYS H H 8.16 0.02 1 120 . 21 CYS HA H 5.063 0.02 1 121 . 21 CYS HB2 H 2.845 0.02 1 122 . 21 CYS HB3 H 2.674 0.02 1 123 . 22 PRO HA H 4.624 0.02 1 124 . 22 PRO HB2 H 2.26 0.02 2 125 . 22 PRO HB3 H 1.938 0.02 2 126 . 22 PRO HG2 H 2.099 0.02 2 127 . 22 PRO HG3 H 2.041 0.02 2 128 . 22 PRO HD2 H 3.742 0.02 2 129 . 22 PRO HD3 H 3.43 0.02 2 130 . 23 GLY H H 8.525 0.02 1 131 . 23 GLY HA2 H 3.838 0.02 1 132 . 23 GLY HA3 H 3.758 0.02 1 133 . 24 ALA H H 8.451 0.02 1 134 . 24 ALA HA H 4.537 0.02 1 135 . 24 ALA HB H 1.366 0.02 1 136 . 25 CYS H H 8.325 0.02 1 137 . 25 CYS HA H 4.59 0.02 1 138 . 25 CYS HB2 H 3.199 0.02 1 139 . 25 CYS HB3 H 3.199 0.02 1 140 . 26 ILE H H 9.083 0.02 1 141 . 26 ILE HA H 4.427 0.02 1 142 . 26 ILE HB H 1.928 0.02 1 143 . 26 ILE HG12 H 1.071 0.02 1 144 . 26 ILE HG13 H 1.071 0.02 1 145 . 26 ILE HG2 H 0.853 0.02 1 146 . 26 ILE HD1 H 0.973 0.02 1 147 . 27 CYS H H 8.951 0.02 1 148 . 27 CYS HA H 4.963 0.02 1 149 . 27 CYS HB2 H 2.811 0.02 1 150 . 27 CYS HB3 H 2.471 0.02 1 151 . 28 ARG H H 8.086 0.02 1 152 . 28 ARG HA H 4.325 0.02 1 153 . 28 ARG HB2 H 2.507 0.02 2 154 . 28 ARG HB3 H 2.067 0.02 2 155 . 28 ARG HG2 H 1.751 0.02 2 156 . 28 ARG HG3 H 1.606 0.02 2 157 . 28 ARG HD2 H 3.254 0.02 1 158 . 28 ARG HD3 H 3.254 0.02 1 159 . 28 ARG HE H 6.99 0.02 2 160 . 29 GLY H H 8.942 0.02 1 161 . 29 GLY HA2 H 3.933 0.02 1 162 . 29 GLY HA3 H 3.933 0.02 1 163 . 30 ASN H H 7.822 0.02 1 164 . 30 ASN HA H 4.706 0.02 1 165 . 30 ASN HB2 H 3.331 0.02 1 166 . 30 ASN HB3 H 2.867 0.02 1 167 . 30 ASN HD21 H 7.59 0.02 2 168 . 30 ASN HD22 H 6.634 0.02 2 169 . 31 GLY H H 8.439 0.02 1 170 . 31 GLY HA2 H 3.999 0.02 2 171 . 31 GLY HA3 H 3.686 0.02 2 172 . 32 TYR H H 7.302 0.02 1 173 . 32 TYR HA H 5.267 0.02 1 174 . 32 TYR HB2 H 3.073 0.02 1 175 . 32 TYR HB3 H 2.652 0.02 1 176 . 32 TYR HD1 H 6.882 0.02 1 177 . 32 TYR HD2 H 6.882 0.02 1 178 . 32 TYR HE1 H 6.707 0.02 1 179 . 32 TYR HE2 H 6.707 0.02 1 180 . 33 CYS H H 8.831 0.02 1 181 . 33 CYS HA H 5.41 0.02 1 182 . 33 CYS HB2 H 3.057 0.02 1 183 . 33 CYS HB3 H 2.801 0.02 1 184 . 34 GLY H H 9.838 0.02 1 185 . 34 GLY HA2 H 4.522 0.02 2 186 . 34 GLY HA3 H 3.967 0.02 2 stop_ save_