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.

scholarly journals Bayesian-Maximum-Entropy reweighting of IDP ensembles based on NMR chemical shifts

2019 ◽  
Author(s):  
Ramon Crehuet ◽  
Pedro J. Buigues ◽  
Xavier Salvatella ◽  
Kresten Lindorff-Larsen
Keyword(s):  
Chemical Shift ◽  
Maximum Entropy ◽  
Chemical Shifts ◽  
Intrinsically Disordered Protein ◽  
Intrinsically Disordered ◽  
Computational Data ◽  
Chemical Shift Data ◽  
Secondary Chemical Shifts ◽  
Validation Set ◽  
Secondary Chemical

AbstractBayesian and Maximum Entropy approaches allow for a statistically sound and systematic fitting of experimental and computational data. Unfortunately, assessing the relative confidence in these two types of data remains difficult as several steps add unknown error. Here we propose the use of a validation-set method to determine the balance, and thus the amount of fitting. We apply the method to synthetic NMR chemical shift data of an intrinsically disordered protein. We show that the method gives consistent results even when other methods to assess the amount of fitting cannot be applied. Finally, we also describe how the errors in the chemical shift predictor can lead to an incorrect fitting and how using secondary chemical shifts could alleviate this problem.

scholarly journals Bayesian-Maximum-Entropy Reweighting of IDP Ensembles Based on NMR Chemical Shifts

Entropy ◽  
2019 ◽  
Vol 21 (9) ◽  
pp. 898 ◽  
Author(s):  
Ramon Crehuet ◽  
Pedro J. Buigues ◽  
Xavier Salvatella ◽  
Kresten Lindorff-Larsen
Keyword(s):  
Chemical Shift ◽  
Maximum Entropy ◽  
Chemical Shifts ◽  
Intrinsically Disordered Protein ◽  
Intrinsically Disordered ◽  
Computational Data ◽  
Chemical Shift Data ◽  
Secondary Chemical Shifts ◽  
Validation Set ◽  
Secondary Chemical

Bayesian and Maximum Entropy approaches allow for a statistically sound and systematic fitting of experimental and computational data. Unfortunately, assessing the relative confidence in these two types of data remains difficult as several steps add unknown error. Here we propose the use of a validation-set method to determine the balance, and thus the amount of fitting. We apply the method to synthetic NMR chemical shift data of an intrinsically disordered protein. We show that the method gives consistent results even when other methods to assess the amount of fitting cannot be applied. Finally, we also describe how the errors in the chemical shift predictor can lead to an incorrect fitting and how using secondary chemical shifts could alleviate this problem.

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.

Immunogold labeling of the calcium binding protein synexin in isolated adrenal chromaffin granules and chromaffin cells

1990 ◽  
Vol 48 (3) ◽  
pp. 952-953
Author(s):  
Gemma A.J. Kuijpers ◽  
Harvey B. Pollard
Keyword(s):  
Adrenal Medulla ◽  
Calcium Binding ◽  
Chromaffin Cells ◽  
Cell Activation ◽  
Structural Information ◽  
Dependent Manner ◽  
Chromaffin Granules ◽  
Immuno Electron Microscopy ◽  
Ultrathin Sections ◽  
Adrenal Chromaffin

Exocytotic fusion of granules in the adrenal medulla chromaffin cell is triggered by a rise in the concentration of cytosolic Ca2+ upon cell activation. The protein synexin, annexin VII, was originally found in the adrenal medulla and has been shown to cause aggregation and to support fusion of chromaffin granules in a Ca2+-dependent manner. We have previously suggested that synexin may there fore play a role in the exocytotic fusion process. In order to obtain more structural information on synexin, we performed immuno-electron microscopy on frozen ultrathin sections of both isolated chromaffin granules and chromaffin cells.Chromaffin granules were isolated from bovine adrenal medulla, and synexin was isolated from bovine lung. Granules were incubated in the presence or absence of synexin (24 μg per mg granule protein) and Ca2+ (1 mM), which induces maximal granule aggregation, in 0.3M sucrose-40m MMES buffer(pH 6.0). Granules were pelleted, washed twice in buffer without synexin and fixed with 2% glutaraldehyde- 2% para formaldehyde in 0.1 M phosphate buffer (GA/PFA) for 30 min. Chromaffin cells were isolated and cultured for 3-5 days, and washed and incubated in Krebs solution with or without 20 uM nicotine. Cells were fixed 90 sec after on set of stimulation with GA/PFA for 30 min. Fixed granule or cell pellets were washed, infiltrated with 2.3 M sucrose in PBS, mounted and frozen in liquid N2.

Scorpion toxin-potassium channel interaction law and its applications.

2020 ◽  
Vol 01 ◽  
Author(s):  
Zheng Zuo ◽  
Zongyun Chen ◽  
Zhijian Cao ◽  
Wenxin Li ◽  
Yingliang Wu
Keyword(s):  
Potassium Channel ◽  
Scorpion Toxin ◽  
Scorpion Toxins ◽  
Toxin Binding ◽  
Channel Interaction ◽  
Binding Interface ◽  
Α Helix ◽  
Interaction Law ◽  
Binding Interfaces ◽  
Β Sheet

: The scorpion toxins are the largest potassium channel-blocking peptide family. The understanding of toxin binding interfaces is usually restricted by two classical binding interfaces: one is the toxin α-helix motif, the other is the antiparallel β-sheet motif. In this review, such traditional knowledge was updated by another two different binding interfaces: one is BmKTX toxin using the turn motif between the α-helix and antiparallel β-sheet domains as the binding interface, the other is Ts toxin using turn motif between the β-sheet in the N-terminal and α-helix domains as the binding interface. Their interaction analysis indicated that the scarce negatively charged residues in the scorpion toxins played a critical role in orientating the toxin binding interface. In view of the toxin negatively charged amino acids as “binding interface regulator”, the law of scorpion toxin-potassium channel interaction was proposed, that is, the polymorphism of negatively charged residue distribution determines the diversity of toxin binding interfaces. Such law was used to develop scorpion toxin-potassium channel recognition control technique. According to this technique, three Kv1.3 channel-targeted peptides, using BmKTX as the template, were designed with the distinct binding interfaces from that of BmKTX through modulating the distribution of toxin negatively charged residues. In view of the potassium channel as the common targets of different animal toxins, the proposed law was also shown to helpfully orientate the binding interfaces of other animal toxins. Clearly, the toxin-potassium channel interaction law would strongly accelerate the research and development of different potassium channelblocking animal toxins in the future.

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