Interaction of troponin I and troponin C: 19F NMR studies of the binding of the inhibitory troponin I peptide to turkey skeletal troponin C

1991 ◽  
Vol 69 (9) ◽  
pp. 674-681 ◽  
Author(s):  
A. Patricia Campbell ◽  
Paul J. Cachia ◽  
Brian D. Sykes
Keyword(s):  
Calcium Binding ◽  
Troponin I ◽  
Dissociation Constants ◽  
Chemical Shifts ◽  
Troponin C ◽  
Resonance Spectroscopy ◽  
Nmr Studies ◽  
Binding Model ◽  
Peptide Binding Site ◽  
Titration Data

We have used 19F nuclear magnetic resonance spectroscopy to study the interaction of the inhibitory region of troponin (TnI) with apo- and calcium(II)-saturated turkey skeletal troponin C (TnC), using the synthetic TnI analogue Nα-acetyl[19FPhe106]TnI(104–115)amide. Dissociation constants of Kd = (3.7 ± 3.1) × 10−5 M for the apo interaction and Kd = (4.8 ± 1.8) × 10−5 M for the calcium(II)-saturated interaction were obtained using a 1:1 binding model of peptide to protein. The 19F NMR chemical shifts for the F-phenylalanine of the bound peptide are different from the apo- and calcium-saturated protein, indicating a different environment for the bound peptide. The possibility of 2:1 binding of the peptide to Ca(II)-saturated TnC was tested by calculating the fit of the experimental titration data to a series of theoretical binding curves in which the dissociation constants for the two hypothetical binding sites were varied. We obtained the best fit for 0.056 mM ≤ Kd1 ≤ 0.071 mM and 0.5 mM ≤ Kd2 ≤ 2.0 mM. These results allow the possibility of a second peptide binding site on calcium(II)-saturated TnC with an affinity 10- to 20-fold weaker than that of the first site.Key words: tropinin C, tropinin I, calcium binding, NMR studies, muscle proteins.

scholarly journals Genomic Organization, Expression, and Analysis of the Troponin C Gene pat-10 of Caenorhabditis elegans

1999 ◽  
Vol 146 (1) ◽  
pp. 193-202 ◽  
Author(s):  
Hiromi Terami ◽  
Benjamin D. Williams ◽  
Shin-ichi Kitamura ◽  
Yasuji Sakube ◽  
Shinji Matsumoto ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Calcium Binding ◽  
Troponin I ◽  
Amino Acid Level ◽  
Troponin C ◽  
In Vivo Studies ◽  
Calcium Binding Site ◽  
Expression Studies ◽  
Missense Alteration

We have cloned and characterized the troponin C gene, pat-10 of the nematode Caenorhabditis elegans. At the amino acid level nematode troponin C is most similar to troponin C of Drosophila (45% identity) and cardiac troponin C of vertebrates. Expression studies demonstrate that this troponin is expressed in body wall muscle throughout the life of the animal. Later, vulval muscles and anal muscles also express this troponin C isoform. The structural gene for this troponin is pat-10 and mutations in this gene lead to animals that arrest as twofold paralyzed embryos late in development. We have sequenced two of the mutations in pat-10 and both had identical two mutations in the gene; one changes D64 to N and the other changes W153 to a termination site. The missense alteration affects a calcium-binding site and eliminates calcium binding, whereas the second mutation eliminates binding to troponin I. These combined biochemical and in vivo studies of mutant animals demonstrate that this troponin is essential for proper muscle function during development.

scholarly journals NMR studies delineating spatial relationships within the cardiac troponin I-troponin C complex.

1994 ◽  
Vol 269 (38) ◽  
pp. 23731-23735
Author(s):  
G.A. Krudy ◽  
Q. Kleerekoper ◽  
X. Guo ◽  
J.W. Howarth ◽  
R.J. Solaro ◽  
...  
Keyword(s):  
Cardiac Troponin ◽  
Troponin I ◽  
Troponin C ◽  
Cardiac Troponin I ◽  
Spatial Relationships ◽  
Nmr Studies ◽  
C Complex

scholarly journals Interact: Automated analysis of protein-ligand interactions by 1D and 2D NMR

2018 ◽  
Author(s):  
Pierre Millard ◽  
Guy Lippens
Keyword(s):  
Dissociation Constants ◽  
Chemical Shifts ◽  
Automated Analysis ◽  
2D Nmr ◽  
Nmr Titration ◽  
Protein Ligand Interactions ◽  
Fitting Procedures ◽  
Ligand Interactions ◽  
Titration Data ◽  
Automated Tools

AbstractNMR titration experiments contain rich information on the thermodynamic, kinetic and structural aspects of protein-ligand interactions. Automated tools are required to process the large number of signals typically acquired in these experiments and facilitate quantitative interpretations. We present Interact, a Python script accessible within the Bruker BioSpin TopSpin™ software, which allows automated analysis of both 1D and 2D NMR titration experiments. Interact performs peak picking and annotation of the successive spectra and supports quantitative interpretation of changes in chemical shifts and linewidths induced by the ligand (e.g. to estimate dissociation constants) through different fitting procedures. Interact can be applied to all types of 1D and 2D NMR experiments and all nuclei, hence facilitating routine analysis of existing and forthcoming NMR titration data. Interact was implemented in Python and can be used on Windows, Unix and MacOS platforms. The source code is distributed under OpenSource license at http://github.com/MetaSys-LISBP/Interact.

Structural information from NMR secondary chemical shifts of peptide α C-H protons in proteins

1983 ◽  
Vol 3 (5) ◽  
pp. 443-452 ◽  
Author(s):  
D. C. Dalgarno ◽  
B. A. Levine ◽  
R. J. P. Williams
Keyword(s):  
Calcium Binding ◽  
Structural Information ◽  
Chemical Shifts ◽  
Troponin C ◽  
Secondary Chemical Shift ◽  
Secondary Chemical Shifts ◽  
Α Helix ◽  
Torsional Angles ◽  
Secondary Chemical ◽  
Β Sheet

The secondary chemical shift experienced by the 1H-NMR resonances of the α C-H protons in proteins can be correlated with their backbone torsional angles ψ, which dictate the orientation of the α C-H proton to the adjacent carbonyl group. It is shown that α C-H protons present in β-sheet regions experience downfield secondary shifts, whereas those in α-helix regions experience upfield secondary shifts. The predictive use of this correlation in assignment studies is illustrated for the calcium-binding protein paravalbumin, for which a crystal structure is available, and troponin C, for which no crystallographic data are available.

1H NMR Conformational Studies in Water of Angiotensin II Analogues Modified at the N- and C-Termini: Interactions of the Aromatic Side Chains and Folding of the N-Terminal Domain

1994 ◽  
Vol 59 (11) ◽  
pp. 2523-2532 ◽  
Author(s):  
John Hondrelis ◽  
John Matsoukas ◽  
George Agelis ◽  
Paul Cordopatis ◽  
Ning Zhou ◽  
...  
Keyword(s):  
Shielding Effect ◽  
Resonance Spectroscopy ◽  
Nmr Studies ◽  
Previous Suggestion ◽  
Aminoisobutyric Acid ◽  
H Nmr ◽  
Neutral Ph ◽  
Angiotensin Ii Analogues ◽  
Proton Resonances ◽  
Cosy Nmr

The conformation of [Sar1]angiotensin II in water at neutral pH has been examined by proton magnetic resonance spectroscopy at 400 MHz and in particular by comparing its 1H NMR spectral data with those of analogues modified at positions 1,4 and 6, namely [Sar1,Cha8]ANGII, [Des Asp1,Cha8]ANGII, [Aib1,Tyr(Me)4]ANGII, [Aib1,Tyr(Me)4,Ile8]ANGII, [N-MeAib1,Tyr(Me)4]ANGII, [N-MeAib1,Tyr(Me)4,Ile8]ANGII, ANGIII and [Sar1,Ile8]ANGII. Assignment of all proton resonances in these analogues was made possible by 2D COSY NMR experiments. The H-2 and H-4 protons for the histidine ring in [Sar1]ANGII, ANGII and ANGIII were shielded compared with the same protons in [Sar1,Ile8]ANGII, [Sar1,Cha8]ANGII and [Des Asp1,Cha8]ANGII; this shielding effect was not disturbed upon methylation of the tyrosine hydroxyl and/or replacement of residue 1 (sarcosine or aspartic acid) with aminoisobutyric acid (Aib) or N-methyl aminoisobutyric acid (N-MeAib). These data are consistent with our previous suggestion based on NMR studies in neutral DMSO that a characteristic folded conformation for ANGII previously observed in non-polar solvents can also be detected in water at neutral pH, but to a lesser degree.

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