scholarly journals Amide proton exchange of a dynamic loop in cell extracts

2013 ◽  
Vol 22 (10) ◽  
pp. 1313-1319 ◽  
Author(s):  
Austin E. Smith ◽  
Mohona Sarkar ◽  
Gregory B. Young ◽  
Gary J. Pielak
Keyword(s):  
Proton Exchange ◽  
Amide Proton ◽  
Amide Proton Exchange ◽  
Cell Extracts

scholarly journals Flexibility of Oxidized and Reduced States of the Chloroplast Regulatory Protein CP12 in Isolation and in Cell Extracts

Biomolecules ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 701
Author(s):  
Helene Launay ◽  
Hui Shao ◽  
Olivier Bornet ◽  
Francois-Xavier Cantrelle ◽  
Regine Lebrun ◽  
...  
Keyword(s):  
Regulatory Protein ◽  
Cell Extract ◽  
Proton Exchange ◽  
Conformational Exchange ◽  
Amide Proton ◽  
Cysteine Residues ◽  
Intrinsically Disordered ◽  
Amide Proton Exchange ◽  
Cell Extracts ◽  
C Terminus

In the chloroplast, Calvin–Benson–Bassham enzymes are active in the reducing environment created in the light by electrons from the photosystems. In the dark, these enzymes are inhibited, mainly caused by oxidation of key regulatory cysteine residues. CP12 is a small protein that plays a role in this regulation with four cysteine residues that undergo a redox transition. Using amide-proton exchange with solvent, measured by nuclear magnetic resonance (NMR) and mass-spectrometry, we confirmed that reduced CP12 is intrinsically disordered. Using real-time NMR, we showed that the oxidation of the two disulfide bridges is simultaneous. In oxidized CP12, the C23–C31 pair is in a region that undergoes a conformational exchange in the NMR-intermediate timescale. The C66–C75 pair is in the C-terminus that folds into a stable helical turn. We confirmed that these structural states exist in a physiologically relevant environment: a cell extract from Chlamydomonas reinhardtii. Consistent with these structural equilibria, the reduction is slower for the C66–C75 pair than for the C23–C31 pair. The redox mid-potentials for the two cysteine pairs differ and are similar to those found for glyceraldehyde 3-phosphate dehydrogenase and phosphoribulokinase, consistent with the regulatory role of CP12.

scholarly journals In-Situ Flexibility of Both Redox-States of the Chloroplast Regulatory Protein CP12

2021 ◽  
Author(s):  
Helene Launay ◽  
Hui Shao ◽  
Olivier Bornet ◽  
Francois-Xavier Cantrelle ◽  
Regine Lebrun ◽  
...  
Keyword(s):  
Regulatory Protein ◽  
Cell Extract ◽  
Proton Exchange ◽  
Conformational Exchange ◽  
Amide Proton ◽  
Cysteine Residues ◽  
Intrinsically Disordered ◽  
Amide Proton Exchange ◽  
C Terminus ◽  
Helical Turn

In the chloroplast, Calvin-Benson-Bassham enzymes are active in the reducing environment imposed in the light via the electrons from the photosystems. In the dark these enzymes are inhibited, and this regulation is mainly mediated via oxidation of key regulatory cysteine residues. CP12 is a small protein that plays a role in this regulation with four cysteine residues that undergo a redox transition. Using amide-proton exchange with solvent measured by nuclear magnetic resonance (NMR) and mass-spectrometry, we confirmed that reduced CP12 is intrinsically disordered. Using real-time NMR, we showed that the oxidation of the two disulfide bridges are simultaneous. In oxidized CP12, the C23-C31 pair is in a region that undergoes a conformational exchange in the NMR-intermediate timescale. The C66-C75 pair is in the C-terminus that folds into a stable helical turn. We confirmed that these structural states exist in a physiologically relevant environment that is, in cell extract from Chlamydomonas reinhardtii. Consistent with these structural equilibria, the reduction is slower for the C66-C75 pair compared to the C23-C31 pair. Finally, the redox mid-potentials for the two cysteine pairs differ and are similar to those found for phosphoribulokinase and glyceraldehyde 3-phosphate dehydrogenase, that we relate to the regulatory role of CP12.

Investigation of Protein Amide-Proton Exchange by 1H Longitudinal Spin Relaxation

1995 ◽  
Vol 108 (3) ◽  
pp. 220-234 ◽  
Author(s):  
Z.W. Zheng ◽  
M.R. Gryk ◽  
M.D. Finucane ◽  
O. Jardetzky
Keyword(s):  
Spin Relaxation ◽  
Proton Exchange ◽  
Amide Proton ◽  
Amide Proton Exchange ◽  
Longitudinal Spin

Corrections - Influence of Charge on the Rate of Amide Proton Exchange

Biochemistry ◽  
1982 ◽  
Vol 21 (12) ◽  
pp. 3036-3036 ◽  
Author(s):  
Peter Kim ◽  
Robert Baldwin
Keyword(s):  
Proton Exchange ◽  
Amide Proton ◽  
Amide Proton Exchange

Effect of peptide binding on amide proton exchange rates in the PDZ2 domain from human phosphatase hPTP1E

1998 ◽  
Vol 76 (2-3) ◽  
pp. 334-340 ◽  
Author(s):  
Irena Ekiel ◽  
Denis Banville ◽  
Shi Hsiang Shen ◽  
Kalle Gehring
Keyword(s):  
Exchange Rates ◽  
Ph Dependence ◽  
Guanidine Hydrochloride ◽  
Peptide Binding ◽  
Fluorescence Emission ◽  
Proton Exchange ◽  
Amide Proton ◽  
Amide Proton Exchange ◽  
C Terminus ◽  
Amide Exchange

Amide hydrogen-deuterium exchange rates were measured in the PDZ2 domain from human phosphatase hPTP1E by 1H-15N heteronuclear NMR spectroscopy. Protection factors were calculated for the slowly exchanging hydrogens in both the free PDZ2 domain and its complex with an octapeptide peptide, R-N-E-I-Q-S-L-V, derived from the C-terminus of the Fas receptor. Aside from a short α-helical region α1 (amino acids A-45 to D-49), the pattern of highly protected amides correlated well with the presence of hydrogen bonds in elements of the secondary structure. Hydrogen-bonded amides showed relatively fast exchange rates with half-lives of less than 9 h at pD 7.6 and 8°C. Protection factors, calculated as the ratio of theoretical (denatured) and observed exchange rates, showed less dispersion in maximal values than did the actual exchange rates. This behavior and the large pH dependence of the exchange rates suggest that amide exchange is close to the EX2 limit. In this limit, exchange of the most protected amides occurs through a global unfolding mechanism. The free energy of the unfolding calculated from the largest protection factors is 4.8 ± 0.4 kcal/mol (1 cal = 4.184 J). This ΔG° closely matches the value measured by experiments with guanidine hydrochloride and fluorescence emission spectroscopy. Peptide binding to PDZ2 resulted in mostly global effects and stabilized the folded domain by 1.4 kcal/mol.Key words: PDZ2 from hPTP1E, amide exchange, ligand binding, NMR.

Sign in / Sign up

Export Citation Format

Share Document