Characterization of the distribution of internal motions in the basic pancreatic trypsin inhibitor using a large number of internal NMR probes

Wagner, Gerhard

doi:10.1017/s0033583500004911

Cited by 274 publications

(210 citation statements)

References 77 publications

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“…In general, amide protons buried in the hydrophobic core or those involved in hydrogen bonding in stable secondary structures are protected from exchange and are observable by NMR after a protein is transferred into deuterated buffer (Wagner, 1983;Englander & Kallenbach, 1983). Those amide protons that exchange most slowly in Pdx in fact correlate well with regions of regular secondary structure (&sheet, a-helices, tight turns) as established by NMR structural methods (Ye et al, 1992).…”

Section: Redox Dependence Of Amide Proton Exchange In Pdxmentioning

confidence: 99%

Redox‐dependent dynamics of putidaredoxin characterized by amide proton exchange

Lyons¹,

Ratnaswamy²,

Pochapsky³

1996

Protein Science

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Multidimensional NMR methods were used to obtain 'H-I'N correlations and "N resonance assignments for amide and side-chain nitrogens of oxidized and reduced putidaredoxin (Pdx), the Fe2S2 ferredoxin, which acts as the physiological reductant of cytochrome P-450,,, (CYPlOl). A model for the solution structure of oxidized Pdx has been determined recently using NMR methods (Pochapsky TC, Ye XM, Ratnaswamy G, Lyons TA, 1994, Biochemistry 33:6424-6432) and redox-dependent ' H NMR spectral features have been described (Pochapsky TC, Ratnaswamy G, Patera A, 1994, Biochemistry33:6433-6441). "N assignments were made with NOESY-('H/''N) HMQC and TOCSY-('H/''N) HSQC spectra obtained using samples of Pdx uniformly labeled with "N. Local dynamics in both oxidation states of Pdx were then characterized by comparison of residue-specific amide proton exchange rates, which were measured by a combination of saturation transfer and H 2 0 / D 2 0 exchange methods at pH 6.4 and 7.4 (uncorrected for isotope effects). In general, where exchange rates for a given site exhibit significant oxidation-state dependence, the oxidized protein exchanges more rapidly than the reduced protein. The largest dependence of exchange rate upon oxidation state is found for residues near the metal center and in a region of compact structure that includes the loop-turn Val 74-Ser 82 and the C-terminal residues (Pro 102-Trp 106). The significance of these findings is discussed in light of the considerable dependence of the binding interaction between Pdx and CYPlOl upon the oxidation state of Pdx.

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Section: Redox Dependence Of Amide Proton Exchange In Pdxmentioning

confidence: 99%

Redox‐dependent dynamics of putidaredoxin characterized by amide proton exchange

Lyons¹,

Ratnaswamy²,

Pochapsky³

1996

Protein Science

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“…Many models describing the protein dynamics that lead to hydrogen exchange in folded proteins have been proposed. These have all been described extensively in several reviews including the breathing model, the penetration model, the mobile defect model, the local unfolding model, and the global unfolding model (Hvidt & Nielsen, 1966;Englander et al, 1972;Englander, 1975: Richards, 1979Wagner & Wuthrich, 1979;Wood-ward & Hilton, 1979;Barksdale & Rosenberg, 1982;Wagner, 1982;Woodward et al, 1982).…”

Section: Hydrogen Exchangementioning

confidence: 99%

Mapping the lifetimes of local opening events in a native state protein

Kragelund¹,

Heinemann²,

Knudsen³

et al. 1998

Protein Science

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The rate constants for the processes that lead to local opening and closing of the structures around hydrogen bonds in native proteins have been determined for most of the secondary structure hydrogen bonds in the four-helix protein acyl coenzyme A binding protein. In an analysis that combines these results with the energies of activation of the opening processes and the stability of the local structures, three groups of residues in the protein structure have been identified. In one group, the structures around the hydrogen bonds have frequent openings, every 600 to 1,500 s, and long lifetimes in the open state, around 1 s. In another group of local structures, the local opening is a very rare event that takes place only every 15 to 60 h. For these the lifetime in the open state is also around 1 s. The majority of local structures have lifetimes between 2,000 and 20,000 s and relatively short lifetimes of the open state in the range between 30 and 400 ms. Mapping of these groups of amides to the tertiary structure shows that the openings of the local structures are not cooperative at native conditions, and they rarely if ever lead to global unfolding. The results suggest a mechanism of hydrogen exchange by progressive local openings.

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“…Analogous solution-state experiments involving two oxygen atoms are even more challenging (CO · · · HO). Assignment of hydroxyl protons is therefore usually obtained via nuclear Overhauser spectroscopy 15 or by analysis of HMBC experiments, 16 which becomes difficult for larger spin systems. † In the solid state, the simultaneous presence of isotropic and anisotropic interactions allows great flexibility in the design of experimental methodologies.…”

Section: Introductionmentioning

confidence: 99%

Identification of Hydroxyl Protons, Determination of Their Exchange Dynamics, and Characterization of Hydrogen Bonding in a Microcrystallin Protein

et al. 2010

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Abstract:Heteronuclear correlation experiments employing perdeuterated proteins enable the observation of all hydroxyl protons in a microcrystalline protein by MAS solid-state NMR. Dipolar-based sequences allow magnetization transfers that are >50 times faster compared to scalar-coupling-based sequences, which significantly facilitates their assignment. Hydroxyl exchange rates were measured using EXSY-type experiments. We find a biexponential decay behavior for those hydroxyl groups that are involved in side chain-side chain C-O-H · · · OdC hydrogen bonds. The quantification of the distances between the hydroxyl proton and the carbon atoms in the hydrogen-bonding donor as well as acceptor group is achieved via a REDOR experiment. In combination with X-ray data and isotropic proton chemical shifts, availability of 1 H, 13 C distance information can aid in the quantitative description of the geometry of these hydrogen bonds. Similarly, correlations between backbone amide proton and carbonyl atoms are observed, which will be useful in the analysis of the registry of -strand arrangement in amyloid fibrils.

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Characterization of the distribution of internal motions in the basic pancreatic trypsin inhibitor using a large number of internal NMR probes

Cited by 274 publications

References 77 publications

Redox‐dependent dynamics of putidaredoxin characterized by amide proton exchange

Redox‐dependent dynamics of putidaredoxin characterized by amide proton exchange

Mapping the lifetimes of local opening events in a native state protein

Identification of Hydroxyl Protons, Determination of Their Exchange Dynamics, and Characterization of Hydrogen Bonding in a Microcrystallin Protein

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