“…If k off ≪ k b , the scheme can be simplified ( , ) as shown in Scheme and or This relaxation would be monoexponential, unlike the previous case, and the dependence of apparent transverse relaxation time on the Na + concentration yields K D , k on , and k off . 2 …”
Section: Resultsmentioning
confidence: 99%
“…cannot be used. However, 23 Na NMR spectroscopy was proven (19)(20)(21)(22)(23) to be a good technique for determining the ion-binding properties of macromolecules under conditions of very fast chemical exchange, and so we chose this method for our study.…”
Section: Resultsmentioning
confidence: 99%
“…For free sodium ions in aqueous solution, where rotation of the ion is very fast, the condition of so-called extreme narrowing, ω 2 τ c 2 , 1 (where ω is the angular Larmor frequency of the nuclei and τ c is the correlation time), is satisfied. In this case, the relaxation of transverse magnetization is characterized by a single-exponential decay with a time constant T 2free that corresponds to a simple Lorentzian line in the 23 Na NMR spectrum with line width ∆ν 1/2 0 where χ is the quadrupolar coupling constant.…”
Section: Resultsmentioning
confidence: 99%
“…Redox Titration of NQR as Monitored by Changes in Line Width of the 23 Na NMR Signal. NQR samples contained 100 µM of the protein in 100 mM KCl, 20 mM Hepes-Tris (pH 7.5), 10% D 2 O, 0.1% DM, and 2 mM NaCl.…”
Section: Methodsmentioning
confidence: 99%
“…
Relaxation characteristics of the 23 Na nuclei magnetization were used to determine the sodiumbinding properties of the Na + -translocating NADH:quinone oxidoreductase from Vibrio harVeyi (NQR). The dissociation constant of Na + for the oxidized enzyme was found to be 24 mM and for the reduced enzyme about 30 µM.
Relaxation characteristics of the 23Na nuclei magnetization were used to determine the sodium-binding properties of the Na+-translocating NADH:quinone oxidoreductase from Vibrio harveyi (NQR). The dissociation constant of Na+ for the oxidized enzyme was found to be 24 mM and for the reduced enzyme about 30 microM. Such large (3 orders in magnitude) redox dependence of the NQR affinity to sodium ions shows that the molecular machinery was designed to use the drop in redox energy for creating an electrochemical sodium gradient. Redox titration of NQR monitored by changes in line width of the 23Na NMR signal at 2 mM Na+ showed that the enzyme affinity to sodium ions follows the Nernst law for a one-electron carrier with Em about -300 mV (vs SHE). The data indicate that energy conservation by NQR involves a mechanism modulating ion affinity by the redox state of an enzyme redox cofactor.
“…If k off ≪ k b , the scheme can be simplified ( , ) as shown in Scheme and or This relaxation would be monoexponential, unlike the previous case, and the dependence of apparent transverse relaxation time on the Na + concentration yields K D , k on , and k off . 2 …”
Section: Resultsmentioning
confidence: 99%
“…cannot be used. However, 23 Na NMR spectroscopy was proven (19)(20)(21)(22)(23) to be a good technique for determining the ion-binding properties of macromolecules under conditions of very fast chemical exchange, and so we chose this method for our study.…”
Section: Resultsmentioning
confidence: 99%
“…For free sodium ions in aqueous solution, where rotation of the ion is very fast, the condition of so-called extreme narrowing, ω 2 τ c 2 , 1 (where ω is the angular Larmor frequency of the nuclei and τ c is the correlation time), is satisfied. In this case, the relaxation of transverse magnetization is characterized by a single-exponential decay with a time constant T 2free that corresponds to a simple Lorentzian line in the 23 Na NMR spectrum with line width ∆ν 1/2 0 where χ is the quadrupolar coupling constant.…”
Section: Resultsmentioning
confidence: 99%
“…Redox Titration of NQR as Monitored by Changes in Line Width of the 23 Na NMR Signal. NQR samples contained 100 µM of the protein in 100 mM KCl, 20 mM Hepes-Tris (pH 7.5), 10% D 2 O, 0.1% DM, and 2 mM NaCl.…”
Section: Methodsmentioning
confidence: 99%
“…
Relaxation characteristics of the 23 Na nuclei magnetization were used to determine the sodiumbinding properties of the Na + -translocating NADH:quinone oxidoreductase from Vibrio harVeyi (NQR). The dissociation constant of Na + for the oxidized enzyme was found to be 24 mM and for the reduced enzyme about 30 µM.
Relaxation characteristics of the 23Na nuclei magnetization were used to determine the sodium-binding properties of the Na+-translocating NADH:quinone oxidoreductase from Vibrio harveyi (NQR). The dissociation constant of Na+ for the oxidized enzyme was found to be 24 mM and for the reduced enzyme about 30 microM. Such large (3 orders in magnitude) redox dependence of the NQR affinity to sodium ions shows that the molecular machinery was designed to use the drop in redox energy for creating an electrochemical sodium gradient. Redox titration of NQR monitored by changes in line width of the 23Na NMR signal at 2 mM Na+ showed that the enzyme affinity to sodium ions follows the Nernst law for a one-electron carrier with Em about -300 mV (vs SHE). The data indicate that energy conservation by NQR involves a mechanism modulating ion affinity by the redox state of an enzyme redox cofactor.
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