data_6656
#######################
# Entry information #
#######################
save_entry_information
_Saveframe_category entry_information
_Entry_title
;
On the Importance of Carbohydrate-Aromatic Interactions for the Molecular
Recognition of Chitooligosaccharides by Hevein Domains. NMR Studies of the
Structure and Binding Affinity of AcAMP2-Like Peptides with non Natural Napthyl
and Fluoroaromatic Residues
;
_BMRB_accession_number 6656
_BMRB_flat_file_name bmr6656.str
_Entry_type original
_Submission_date 2005-06-06
_Accession_date 2005-06-06
_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 Chavez M. Isabel .
2 Andreu Cecilia . .
3 Vidal Paloma . .
4 Freire Felix . .
5 Aboitiz Nuria . .
6 Groves Patrick . .
7 Asensio Juan L. .
8 Asensio Gregorio . .
9 Muraki Michiro . .
10 Canada F. Javier .
11 Jimenez-Barbero Jesus . .
stop_
loop_
_Saveframe_category_type
_Saveframe_category_type_count
assigned_chemical_shifts 1
stop_
loop_
_Data_type
_Data_type_count
"1H chemical shifts" 142
stop_
loop_
_Revision_date
_Revision_keyword
_Revision_author
_Revision_detail
2008-07-31 update BMRB 'updating non-standard residues'
2005-12-22 original author 'Original release'
stop_
loop_
_Related_BMRB_accession_number
_Relationship
6591 AcAMP2F18Pff/Y20Pff
6637 'AcAMP2F18Nalb mutant'
6639 'AcAMP2F18W mutant'
6647 AcAMP2F18Wb
6657 'AcAMP2F18Nal mutant'
stop_
save_
#############################
# Citation for this entry #
#############################
save_entry_citation
_Saveframe_category entry_citation
_Citation_full .
_Citation_title
;
On the Importance of Carbohydrate-Aromatic Interactions for the Molecular
Recognition of Chitooligosaccharides by Hevein Domains. NMR Studies of the
Structure and Binding Affinity of AcAMP2-Like Peptides with non Natural Napthyl
and Fluoroaromatic Residues
;
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 16220560
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Chavez M. Isabel .
2 Andreu Cecilia . .
3 Vidal Paloma . .
4 Aboitiz Nuria . .
5 Freire Felix . .
6 Groves Patrick . .
7 Asensio Juan L. .
8 Asensio Gregorio . .
9 Muraki Michiro . .
10 Canada F. Javier .
11 Jimenez-Barbero Jesus . .
stop_
_Journal_abbreviation 'Chem. Eur. J.'
_Journal_volume 11
_Journal_issue 23
_Journal_CSD .
_Book_chapter_title .
_Book_volume .
_Book_series .
_Book_ISBN .
_Conference_state_province .
_Conference_abstract_number .
_Page_first 7060
_Page_last 7074
_Year 2005
_Details .
loop_
_Keyword
AcAMP2
'carbohydrate binding'
Chitin
hevein
'Molecular Dynamics'
'molecular recognition'
NMR
protein
stop_
save_
#######################################
# Cited references within the entry #
#######################################
save_reference-1
_Saveframe_category citation
_Citation_full
;
Martins JC, Maes D, Loris R, Pepermans HA, Wyns L, Willem R, Verheyden P.
H NMR study of the solution structure of Ac-AMP2, a sugar binding
antimicrobial protein isolated from Amaranthus caudatus. J Mol Biol. 1996 May
3;258(2):322-33.
;
_Citation_title 'H NMR study of the solution structure of Ac-AMP2, a sugar binding antimicrobial protein isolated from Amaranthus caudatus.'
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 8627629
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Martins 'J. C.' C. .
2 Maes D. . .
3 Loris R. . .
4 Pepermans 'H. A.' A. .
5 Wyns L. . .
6 Willem R. . .
7 Verheyden P. . .
stop_
_Journal_abbreviation 'J. Mol. Biol.'
_Journal_name_full 'Journal of molecular biology'
_Journal_volume 258
_Journal_issue 2
_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 322
_Page_last 333
_Year 1996
_Details
;
The conformation in water of antimicrobial protein 2 from Amaranthus caudatus
(Ac-AMP2) was determined using 1H NMR, DIANA and restrained molecular modeling.
Ac-AMP2 is a 30 amino acid residue, lectin-like protein that specifically binds
to chitin, a polymer of beta-1,4-N-acetyl-D-glucosamine. After sequence
specific resonance assignments, a total of 198 distance restraints were
collected from 2D NOESY buildup spectra at 500 MHz at pH 2, supplemented by a
2D NOESY spectrum at 600 MHz. The location of the three previously unassigned
disulfide bridges was determined from preliminary DIANA structures, using a
statistical analysis of intercystinyl distances. The solution structure of
Ac-AMP2 is presented as a set of 26 DIANA structures, further refined by
restrained molecular dynamics using a simulated annealing protocol in the AMBER
force field, with a backbone r.m.s.d. for the well defined Glu3-Cys28 segment
of 0.69(+/-0.12) angstroms. The main structural element is an antiparallel
beta-sheet from Met13 to Lys23 including a betaI-turn over Gln17-Phel8 with a
beta bulge at Gly19. In addition, a beta'I turn over Arg6-Gly7, a beta'III turn
over Ser11-Gly12 and a helical turn from Gly24 to Cys28 are identified. This
structure is very similar to the equivalent regions of the X-ray structure of
wheat germ agglutinin and the NMR structure of hevein.
;
save_
save_reference-2
_Saveframe_category citation
_Citation_full
;
Muraki M.
The importance of CH/pi interactions to the function of carbohydrate binding
proteins. Protein Pept Lett. 2002 Jun;9(3):195-209. Review.
;
_Citation_title 'The importance of CH/pi interactions to the function of carbohydrate binding proteins.'
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 12144516
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Muraki Michiro . .
stop_
_Journal_abbreviation 'Protein Pept. Lett.'
_Journal_name_full 'Protein and peptide letters'
_Journal_volume 9
_Journal_issue 3
_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 195
_Page_last 209
_Year 2002
_Details
;
It is suggested that the interactions between the hydrophobic C-H groups of
carbohydrate residues and the pi-electron systems of aromatic amino-acid
residues play an important role in the ligand-recognition function of
carbohydrate-binding proteins. This review focuses on our recent structural and
functional studies of human lysozyme and hevein-domain type lectins (wheat-germ
agglutinin and Ac-AMP2) aimed at understanding how CH/pi interactions are
involved in the actual binding events.
;
save_
save_reference-3
_Saveframe_category citation
_Citation_full
;
Aboitiz N, Vila-Perello M, Groves P, Asensio JL, Andreu D, Canada FJ,
Jimenez-Barbero J.
NMR and modeling studies of protein-carbohydrate interactions: synthesis,
three-dimensional structure, and recognition properties of a minimum hevein
domain with binding affinity for chitooligosaccharides.
Chembiochem. 2004 Sep 6;5(9):1245-55.
;
_Citation_title 'NMR and modeling studies of protein-carbohydrate interactions: synthesis, three-dimensional structure, and recognition properties of a minimum hevein domain with binding affinity for chitooligosaccharides.'
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 15368576
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Aboitiz Nuria . .
2 Vila-Perello Miquel . .
3 Groves Patrick . .
4 Asensio 'Juan Luis' L. .
5 Andreu David . .
6 Canada 'Francisco Javier' J. .
7 Jimenez-Barbero Jesus . .
stop_
_Journal_abbreviation Chembiochem
_Journal_name_full 'Chembiochem : a European journal of chemical biology'
_Journal_volume 5
_Journal_issue 9
_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 1245
_Page_last 1255
_Year 2004
_Details
;
HEV32, a 32-residue, truncated hevein lacking eleven C-terminal amino acids, was
synthesized by solid-phase methodology and correctly folded with three cysteine
bridge pairs. The affinities of HEV32 for small chitin fragments--in the forms
of N,N',N"-triacetylchitotriose ((GlcNAc)3) (millimolar) and
N,N',N",N"',N"",N""'-hexaacetylchitohexaose ((GlcNAc)6) (micromolar)--as
measured by NMR and fluorescence methods, are comparable with those of native
hevein. The HEV32 ligand-binding process is enthalpy driven, while entropy
opposes binding. The NMR structure of ligand-bound HEV32 in aqueous solution
was determined to be highly similar to the NMR structure of ligand-bound
hevein. Solvated molecular-dynamics simulations were performed in order to
monitor the changes in side-chain conformation of the binding site of HEV32 and
hevein upon interaction with ligands. The calculations suggest that the Trp21
side-chain orientation of HEV32 in the free form differs from that in the bound
state; this agrees with fluorescence and thermodynamic data. HEV32 provides a
simple molecular model for studying protein-carbohydrate interactions and for
understanding the physiological relevance of small native hevein domains
lacking C-terminal residues.
;
save_
save_reference-4
_Saveframe_category citation
_Citation_full
;
Asensio JL, Siebert HC, von Der Lieth CW, Laynez J, Bruix M, Soedjanaamadja
UM, Beintema JJ, Canada FJ, Gabius HJ, Jimenez-Barbero J.
NMR investigations of protein-carbohydrate interactions: studies on the
relevance of Trp/Tyr variations in lectin binding sites as deduced from
titration microcalorimetry and NMR studies on hevein domains. Determination of
the NMR structure of the complex between pseudohevein and
N,N',N"-triacetylchitotriose. Proteins. 2000 Aug 1;40(2):218-36.
;
_Citation_title 'NMR investigations of protein-carbohydrate interactions: studies on the relevance of Trp/Tyr variations in lectin binding sites as deduced from titration microcalorimetry and NMR studies on hevein domains. Determination of the NMR structure of the complex between pseudohevein and N,N',N"-triacetylchitotriose.'
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 10842338
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Asensio 'J. L.' L. .
2 Siebert 'H. C.' C. .
3 'von Der Lieth' 'C. W.' W. .
4 Laynez J. . .
5 Bruix M. . .
6 Soedjanaamadja 'U. M.' M. .
7 Beintema 'J. J.' J. .
8 Canada 'F. J.' J. .
9 Gabius 'H. J.' J. .
10 Jimenez-Barbero J. . .
stop_
_Journal_abbreviation Proteins
_Journal_name_full Proteins
_Journal_volume 40
_Journal_issue 2
_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 218
_Page_last 236
_Year 2000
_Details
;
Model studies on lectins and their interactions with carbohydrate ligands in
solution are essential to gain insights into the driving forces for complex
formation and to optimize programs for computer simulations. The specific
interaction of pseudohevein with N,N', N"-triacetylchitotriose has been
analyzed by (1)H-NMR spectroscopy. Because of its small size, with a chain
length of 45 amino acids, this lectin is a prime target to solution-structure
determination by NOESY NMR experiments in water. The NMR-analysis was extended
to assessment of the topology of the complex between pseudohevein and N,
N',N"-triacetylchitotriose. NOESY experiments in water solution provided 342
protein proton-proton distance constraints. Binding of the ligand did not
affect the pattern of the protein nuclear Overhauser effect signal noticeably,
what would otherwise be indicative of a ligand-induced conformational change.
The average backbone (residues 3-41) RMSD of the 20 refined structures was 1.14
A, whereas the heavy atom RMSD was 2.18 A. Two different orientations of the
trisaccharide within the pseudohevein binding site are suggested, furnishing an
explanation in structural terms for the lectin's capacity to target chitin. In
both cases, hydrogen bonds and van der Waals contacts confer stability to the
complexes. This conclusion is corroborated by the thermodynamic parameters of
binding determined by NMR and isothermal titration calorimetry. The association
process was enthalpically driven. In relation to hevein, the
Trp/Tyr-substitution in the binding pocket has only a small effect on the free
energy of binding in contrast to engineered galectin-1 and a mammalian C-type
lectin. A comparison of the three-dimensional structure of pseudohevein in
solution to those reported for wheat germ agglutinin (WGA) in the solid state
and for hevein and WGA-B in solution has been performed, providing a data
source about structural variability of the hevein domains. The experimentally
derived structures and the values of the solvent accessibilities for several
key residues have also been compared with conformations obtained by molecular
dynamics simulations, pointing to the necessity to further refine the programs
to enhance their predictive reliability and, thus, underscoring the importance
of this kind of combined analysis in model systems.
;
save_
save_reference-5
_Saveframe_category citation
_Citation_full
;
Asensio JL, Canada FJ, Siebert HC, Laynez J, Poveda A, Nieto PM,
Soedjanaamadja UM, Gabius HJ, Jimenez-Barbero J.
Structural basis for chitin recognition by defense proteins: GlcNAc residues
are bound in a multivalent fashion by extended binding sites in hevein
domains. Chem Biol. 2000 Jul;7(7):529-43.
;
_Citation_title 'Structural basis for chitin recognition by defense proteins: GlcNAc residues are bound in a multivalent fashion by extended binding sites in hevein domains.'
_Citation_status published
_Citation_type journal
_CAS_abstract_code .
_MEDLINE_UI_code .
_PubMed_ID 10903932
loop_
_Author_ordinal
_Author_family_name
_Author_given_name
_Author_middle_initials
_Author_family_title
1 Asensio 'J. L.' L. .
2 Canada 'F. J.' J. .
3 Siebert 'H. C.' C. .
4 Laynez J. . .
5 Poveda A. . .
6 Nieto 'P. M.' M. .
7 Soedjanaamadja 'U. M.' M. .
8 Gabius 'H. J.' J. .
9 Jimenez-Barbero J. . .
stop_
_Journal_abbreviation 'Chem. Biol.'
_Journal_name_full 'Chemistry & biology'
_Journal_volume 7
_Journal_issue 7
_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 529
_Page_last 543
_Year 2000
_Details
;
BACKGROUND: Many plants respond to pathogenic attack by producing defense
proteins that are capable of reversible binding to chitin, a polysaccharide
present in the cell wall of fungi and the exoskeleton of insects. Most of these
chitin-binding proteins include a common structural motif of 30 to 43 residues
organized around a conserved four-disulfide core, known as the 'hevein domain'
or 'chitin-binding' motif. Although a number of structural and thermodynamic
studies on hevein-type domains have been reported, these studies do not clarify
how chitin recognition is achieved. RESULTS: The specific interaction of hevein
with several (GlcNAc)(n) oligomers has been studied using nuclear magnetic
resonance (NMR), analytical ultracentrifugation and isothermal titration
microcalorimetry (ITC). The data demonstrate that hevein binds (GlcNAc)(2-4) in
1:1 stoichiometry with millimolar affinity. In contrast, for (GlcNAc)(5), a
significant increase in binding affinity is observed. Analytical
ultracentrifugation studies on the hevein-(GlcNAc)(5,8) interaction allowed
detection of protein-carbohydrate complexes with a ratio of 2:1 in solution.
NMR structural studies on the hevein-(GlcNAc)(5) complex showed the existence
of an extended binding site with at least five GlcNAc units directly involved
in protein-sugar contacts. CONCLUSIONS: The first detailed structural model for
the hevein-chitin complex is presented on the basis of the analysis of NMR
data. The resulting model, in combination with ITC and analytical
ultracentrifugation data, conclusively shows that recognition of chitin by
hevein domains is a dynamic process, which is not exclusively restricted to the
binding of the nonreducing end of the polymer as previously thought. This
allows chitin to bind with high affinity to a variable number of protein
molecules, depending on the polysaccharide chain length. The biological process
is multivalent.
;
save_
##################################
# Molecular system description #
##################################
save_system_AcAMP2F18Pff_F20Pfff
_Saveframe_category molecular_system
_Mol_system_name 'AcAMP2F18Pff/F20Pfff mutant'
_Abbreviation_common AcAMP2F18Pff/F20Pfff
_Enzyme_commission_number .
loop_
_Mol_system_component_name
_Mol_label
AcAMP2F18Pff/F20Pfff $AcAMP2F18Pff_F20Pfff
stop_
_System_molecular_weight .
_System_physical_state native
_System_oligomer_state monomer
_System_paramagnetic no
_System_thiol_state 'all disulfide bound'
loop_
_Biological_function
'chitin binding lectin'
stop_
_Database_query_date .
_Details .
save_
########################
# Monomeric polymers #
########################
save_AcAMP2F18Pff_F20Pfff
_Saveframe_category monomeric_polymer
_Mol_type polymer
_Mol_polymer_class protein
_Name_common AcAMP2
_Name_variant 'AcAMP2F18Pff/Y20Pfff mutant'
_Abbreviation_common AcAMP2F18Pff/Y20Pfff
_Molecular_mass 3209
_Mol_thiol_state 'all disulfide bound'
_Details
;
The secondary structure of this
protein presents beta-sheet on residues over residues 13 to 23.
This protein is a hevein domain, and has high homology with AcAMP2
antimicrobial peptide. Carbohydrate binding site involves residues S16, F18FF,
F20FF and Y27.
;
##############################
# Polymer residue sequence #
##############################
_Residue_count 30
_Mol_residue_sequence
;
VGECVRGRCPSGMCCSQXGX
CGKGPKYCGR
;
loop_
_Residue_seq_code
_Residue_label
1 VAL 2 GLY 3 GLU 4 CYS 5 VAL
6 ARG 7 GLY 8 ARG 9 CYS 10 PRO
11 SER 12 GLY 13 MET 14 CYS 15 CYS
16 SER 17 GLN 18 PFF 19 GLY 20 PFF
21 CYS 22 GLY 23 LYS 24 GLY 25 PRO
26 LYS 27 TYR 28 CYS 29 GLY 30 ARG
stop_
_Sequence_homology_query_date .
_Sequence_homology_query_revised_last_date 2015-01-28
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 6591 AcAMP2F18Pff/Y20Pff 100.00 30 100.00 100.00 1.34e-08
PDB 1ZNT "18 Nmr Structures Of Acamp2-Like Peptide With Non Natural Fluoroaromatic Residue (Acamp2f18pffY20PFF) COMPLEX WITH N,N,N- Triac" 100.00 31 100.00 100.00 1.35e-08
stop_
save_
######################
# Polymer residues #
######################
save_chem_comp_PFF
_Saveframe_category polymer_residue
_Mol_type 'L-peptide linking'
_Name_common 4-FLUORO-L-PHENYLALANINE
_BMRB_code .
_PDB_code PFF
_Standard_residue_derivative .
_Molecular_mass 183.180
_Mol_paramagnetic .
_Details
;
Information obtained from PDB's Chemical Component Dictionary
at http://wwpdb-remediation.rutgers.edu/downloads.html
Downloaded on Mon Aug 8 14:18:20 2011
;
loop_
_Atom_name
_PDB_atom_name
_Atom_type
_Atom_chirality
_Atom_charge
_Atom_oxidation_number
_Atom_unpaired_electrons
N N N . 0 . ?
CA CA C . 0 . ?
C C C . 0 . ?
O O O . 0 . ?
OXT OXT O . 0 . ?
CB CB C . 0 . ?
CG CG C . 0 . ?
CD1 CD1 C . 0 . ?
CD2 CD2 C . 0 . ?
CE1 CE1 C . 0 . ?
CE2 CE2 C . 0 . ?
CZ CZ C . 0 . ?
F F F . 0 . ?
H H H . 0 . ?
H2 H2 H . 0 . ?
HA HA H . 0 . ?
HXT HXT H . 0 . ?
HB2 HB2 H . 0 . ?
HB3 HB3 H . 0 . ?
HD1 HD1 H . 0 . ?
HD2 HD2 H . 0 . ?
HE1 HE1 H . 0 . ?
HE2 HE2 H . 0 . ?
stop_
loop_
_Bond_order
_Bond_atom_one_atom_name
_Bond_atom_two_atom_name
_PDB_bond_atom_one_atom_name
_PDB_bond_atom_two_atom_name
SING N CA ? ?
SING N H ? ?
SING N H2 ? ?
SING CA C ? ?
SING CA CB ? ?
SING CA HA ? ?
DOUB C O ? ?
SING C OXT ? ?
SING OXT HXT ? ?
SING CB CG ? ?
SING CB HB2 ? ?
SING CB HB3 ? ?
DOUB CG CD1 ? ?
SING CG CD2 ? ?
SING CD1 CE1 ? ?
SING CD1 HD1 ? ?
DOUB CD2 CE2 ? ?
SING CD2 HD2 ? ?
DOUB CE1 CZ ? ?
SING CE1 HE1 ? ?
SING CE2 CZ ? ?
SING CE2 HE2 ? ?
SING CZ F ? ?
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
$AcAMP2F18Pff_F20Pfff 'inca wheat' 3567 Eukaryota Viridiplantae Amaranthus caudatus 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
_Details
$AcAMP2F18Pff_F20Pfff 'chemical synthesis' . . . . .
;
The residues Phe18 and Tyr20 of the native AcAMP2 were changed to
4-fluorophenylalanine. The aminoacids were manually assembled by solid phase
synthesis using Fmoc chemistry according to satandard protocols.
;
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
$AcAMP2F18Pff_F20Pfff 2.0 mM .
H2O 90 % .
D2O 10 % .
stop_
save_
############################
# Computer software used #
############################
save_XWINNMR
_Saveframe_category software
_Name xwinnmr
_Version 3.2
loop_
_Task
'Data collection'
stop_
_Details 'The software performs acquisition and processing of NMR experiments'
save_
save_XEASY
_Saveframe_category software
_Name XEASY
_Version 1.3.13
loop_
_Task
'Data analysis'
stop_
_Details
;
Bartels C., Xia T., Billeter M., Guntert P. and Wuthrich K. (1995) J. Biol. NMR
6, 1-10. The program XEASY for computer-supported NMR spectral analysis of
biological macromolecules.
This program helps visualization and assignment of 2D NMR spectra.
;
save_
save_DYANA
_Saveframe_category software
_Name DYANA
_Version 1.5
loop_
_Task
'Structure solution'
stop_
_Details
;
Guntert P., Mumenthaler C. and Wuthrich K. (1997) J. Mol. Biol. 273, 283-298.
Torsion angle dynamics for NMR structure calculation with the new program
DYANA.
This program yields a collection of protein structures that fit the 1H-1H
distance constraints experimentally obtained.
;
save_
save_AMBER
_Saveframe_category software
_Name AMBER
_Version 5.0
loop_
_Task
'Structure refinement'
stop_
_Details
;
Pearlman D. A., Case D. A., Caldwell J. W., Cheatham T. E., DeBolt S., Ross W.
S., Ferguson D., Seibel G. L., and Kollman P. A. (1995) Comp. Phys. Commun. 91,
1-41. AMBER, a package of computer programs for applying molecular mechanics,
normal mode analysis, molecular dynamics and free energy calculations to
simulate the structural and energetic properties of molecules."
The programs used from this package perform molecular dynamics calculations on
protein structures. They also convert dyana-format pdb files into amber-format
pdb files.
;
save_
#########################
# Experimental detail #
#########################
##################################
# NMR Spectrometer definitions #
##################################
save_NMR_spectrometer
_Saveframe_category NMR_spectrometer
_Manufacturer Bruker
_Model AMX
_Field_strength 500
_Details .
save_
#############################
# NMR applied experiments #
#############################
save_TOCSY_1
_Saveframe_category NMR_applied_experiment
_Experiment_name TOCSY
_Sample_label $sample-1
save_
save_NOESY_2
_Saveframe_category NMR_applied_experiment
_Experiment_name NOESY
_Sample_label $sample-1
save_
save_NMR_applied_experiment
_Saveframe_category NMR_applied_experiment
_Experiment_name .
_BMRB_pulse_sequence_accession_number .
_Details .
save_
save_NMR_spec_expt__0_1
_Saveframe_category NMR_applied_experiment
_Experiment_name TOCSY
_BMRB_pulse_sequence_accession_number .
_Details .
save_
save_NMR_spec_expt__0_2
_Saveframe_category NMR_applied_experiment
_Experiment_name NOESY
_BMRB_pulse_sequence_accession_number .
_Details .
save_
#######################
# Sample conditions #
#######################
save_conditions-1
_Saveframe_category sample_conditions
_Details .
loop_
_Variable_type
_Variable_value
_Variable_value_error
_Variable_value_units
'ionic strength' 0.15 0.02 M
pH 5.6 0.2 pH
temperature 298 0.2 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
DSS H 2 'methyl protons' ppm 0.0 . indirect . . . 0.153506088
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_AcAMP2F18Pff_Y20Pfff_shifts
_Saveframe_category assigned_chemical_shifts
_Details
;
The atom HA2 of GLY22 shows up at 1.828; due the proximity to the aromatic
system of the residue 27
;
loop_
_Sample_label
$sample-1
stop_
_Sample_conditions_label $conditions-1
_Chem_shift_reference_set_label $chemical_shift_reference
_Mol_system_component_name AcAMP2F18Pff/F20Pfff
_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 VAL HA H 3.831 0.004 1
2 . 1 VAL HB H 2.143 0.007 1
3 . 1 VAL HG1 H 0.989 0.005 2
4 . 1 VAL HG2 H 1.038 0.02 2
5 . 2 GLY H H 8.969 0.02 1
6 . 2 GLY HA2 H 3.836 0.001 2
7 . 2 GLY HA3 H 4.224 0.012 2
8 . 3 GLU H H 9.095 0.003 1
9 . 3 GLU HA H 4.255 0.006 1
10 . 3 GLU HB2 H 1.902 0.005 2
11 . 3 GLU HB3 H 1.952 0.008 2
12 . 3 GLU HG2 H 2.362 0.007 2
13 . 4 CYS H H 7.654 0.001 1
14 . 4 CYS HA H 4.420 0.005 1
15 . 4 CYS HB2 H 2.770 0.005 2
16 . 4 CYS HB3 H 3.080 0.004 2
17 . 5 VAL H H 8.398 0.002 1
18 . 5 VAL HA H 4.069 0.003 1
19 . 5 VAL HB H 1.868 0.015 1
20 . 5 VAL HG1 H 0.841 0.007 2
21 . 6 ARG H H 9.381 0.003 1
22 . 6 ARG HA H 3.828 0.003 1
23 . 6 ARG HB2 H 1.807 0.008 2
24 . 6 ARG HB3 H 1.885 0.006 2
25 . 6 ARG HG2 H 1.557 0.008 2
26 . 6 ARG HD2 H 3.196 0.007 2
27 . 6 ARG HE H 7.145 0.001 1
28 . 7 GLY H H 8.277 0.002 1
29 . 7 GLY HA2 H 3.679 0.02 2
30 . 7 GLY HA3 H 4.064 0.001 2
31 . 8 ARG H H 7.837 0.004 1
32 . 8 ARG HA H 4.631 0.006 1
33 . 8 ARG HB2 H 1.807 0.006 2
34 . 8 ARG HB3 H 1.857 0.001 2
35 . 8 ARG HG2 H 1.538 0.006 2
36 . 8 ARG HG3 H 1.630 0.004 2
37 . 8 ARG HD2 H 3.143 0.002 2
38 . 8 ARG HH21 H 6.781 0.001 2
39 . 8 ARG HH22 H 7.339 0.001 2
40 . 9 CYS H H 8.608 0.005 1
41 . 9 CYS HA H 5.190 0.003 1
42 . 9 CYS HB2 H 2.225 0.007 2
43 . 9 CYS HB3 H 2.813 0.010 2
44 . 10 PRO HA H 4.405 0.003 1
45 . 10 PRO HB2 H 1.672 0.009 2
46 . 10 PRO HB3 H 2.340 0.006 2
47 . 10 PRO HG2 H 1.911 0.005 2
48 . 10 PRO HG3 H 1.960 0.007 2
49 . 10 PRO HD2 H 3.433 0.007 2
50 . 10 PRO HD3 H 3.916 0.004 2
51 . 11 SER H H 8.278 0.002 1
52 . 11 SER HA H 4.105 0.004 1
53 . 11 SER HB2 H 3.677 0.02 2
54 . 12 GLY H H 8.876 0.002 1
55 . 12 GLY HA2 H 3.627 0.004 2
56 . 12 GLY HA3 H 4.188 0.02 2
57 . 13 MET H H 7.907 0.002 1
58 . 13 MET HA H 4.637 0.009 1
59 . 13 MET HB2 H 1.717 0.006 2
60 . 13 MET HB3 H 1.985 0.006 2
61 . 13 MET HG2 H 2.287 0.006 2
62 . 14 CYS H H 9.212 0.005 1
63 . 14 CYS HA H 4.568 0.007 1
64 . 14 CYS HB2 H 2.360 0.004 2
65 . 14 CYS HB3 H 3.801 0.001 2
66 . 15 CYS H H 8.769 0.003 1
67 . 15 CYS HA H 4.727 0.02 1
68 . 15 CYS HB2 H 2.879 0.002 2
69 . 16 SER HA H 4.893 0.001 1
70 . 16 SER HB2 H 4.376 0.004 2
71 . 17 GLN H H 9.133 0.003 1
72 . 17 GLN HA H 4.027 0.008 1
73 . 17 GLN HB2 H 1.655 0.002 2
74 . 17 GLN HB3 H 1.811 0.006 2
75 . 17 GLN HG3 H 1.573 0.004 2
76 . 17 GLN HE21 H 7.041 0.007 2
77 . 17 GLN HE22 H 7.588 0.02 2
78 . 18 PFF H H 7.489 0.007 1
79 . 18 PFF HA H 4.714 0.02 1
80 . 18 PFF HB2 H 2.791 0.003 2
81 . 18 PFF HB3 H 3.614 0.006 2
82 . 18 PFF HD1 H 7.254 0.002 3
83 . 18 PFF HE1 H 7.048 0.02 3
84 . 19 GLY H H 7.850 0.001 1
85 . 19 GLY HA2 H 3.530 0.002 2
86 . 19 GLY HA3 H 3.925 0.005 2
87 . 20 PFF H H 7.517 0.006 1
88 . 20 PFF HA H 5.064 0.007 1
89 . 20 PFF HB2 H 2.884 0.006 2
90 . 20 PFF HB3 H 3.342 0.006 2
91 . 20 PFF HD1 H 7.035 0.004 3
92 . 20 PFF HE1 H 6.680 0.005 3
93 . 21 CYS H H 8.919 0.004 1
94 . 21 CYS HA H 5.503 0.002 1
95 . 21 CYS HB2 H 2.799 0.02 2
96 . 22 GLY H H 8.422 0.004 1
97 . 22 GLY HA2 H 1.828 0.003 2
98 . 22 GLY HA3 H 3.611 0.005 2
99 . 23 LYS H H 8.034 0.004 1
100 . 23 LYS HA H 4.899 0.005 1
101 . 23 LYS HB2 H 1.621 0.008 2
102 . 23 LYS HB3 H 1.825 0.006 2
103 . 23 LYS HG2 H 1.309 0.007 2
104 . 23 LYS HG3 H 1.403 0.004 2
105 . 23 LYS HD2 H 1.697 0.02 2
106 . 23 LYS HE2 H 2.850 0.02 2
107 . 24 GLY H H 8.347 0.003 1
108 . 24 GLY HA2 H 3.917 0.002 2
109 . 24 GLY HA3 H 4.538 0.005 2
110 . 25 PRO HA H 4.251 0.005 1
111 . 25 PRO HB2 H 1.908 0.003 2
112 . 25 PRO HG2 H 2.021 0.005 2
113 . 25 PRO HG3 H 2.272 0.004 2
114 . 25 PRO HD2 H 3.606 0.001 2
115 . 25 PRO HD3 H 3.809 0.006 2
116 . 26 LYS H H 8.780 0.002 1
117 . 26 LYS HA H 3.976 0.004 1
118 . 26 LYS HB2 H 1.544 0.011 2
119 . 26 LYS HB3 H 1.744 0.005 2
120 . 26 LYS HG2 H 1.075 0.007 2
121 . 26 LYS HG3 H 1.343 0.004 2
122 . 27 TYR H H 7.609 0.002 1
123 . 27 TYR HA H 4.084 0.004 1
124 . 27 TYR HB2 H 2.476 0.006 2
125 . 27 TYR HB3 H 2.931 0.004 2
126 . 27 TYR HD1 H 7.143 0.002 3
127 . 27 TYR HE1 H 6.683 0.007 3
128 . 28 CYS H H 8.465 0.001 1
129 . 28 CYS HA H 4.519 0.009 1
130 . 28 CYS HB2 H 2.712 0.007 2
131 . 28 CYS HB3 H 3.205 0.006 2
132 . 29 GLY H H 7.975 0.002 1
133 . 29 GLY HA2 H 3.973 0.010 2
134 . 30 ARG H H 8.273 0.002 1
135 . 30 ARG HA H 4.261 0.008 1
136 . 30 ARG HB2 H 1.682 0.002 2
137 . 30 ARG HB3 H 1.838 0.004 2
138 . 30 ARG HG2 H 1.547 0.003 2
139 . 30 ARG HD2 H 3.147 0.004 2
140 . 30 ARG HE H 7.145 0.001 1
141 . 30 ARG HH21 H 7.011 0.02 1
142 . 30 ARG HH22 H 7.011 0.02 1
stop_
save_