Apparatus for steady-state kinetic measurements of the catalytic reduction of nitric oxide by ammonia on iron oxide

1991 ◽  
Vol 14 (1) ◽  
pp. 39-44 ◽  
Author(s):  
Michael Claußen ◽  
Hartmut Kainer ◽  
Ulrich Kunz ◽  
Ulrich Hoffmann
Keyword(s):  
Nitric Oxide ◽  
Steady State ◽  
Iron Oxide ◽  
Catalytic Reduction ◽  
Kinetic Measurements ◽  
Steady State Kinetic ◽  
State Kinetic

scholarly journals Dihydrolipoamide dehydrogenase from halophilic archaebacteria

1984 ◽  
Vol 218 (3) ◽  
pp. 811-818 ◽  
Author(s):  
M J Danson ◽  
R Eisenthal ◽  
S Hall ◽  
S R Kessell ◽  
D L Williams
Keyword(s):  
Steady State ◽  
Iodoacetic Acid ◽  
Enzymic Activity ◽  
Evolutionary Significance ◽  
Dihydrolipoamide Dehydrogenase ◽  
Kinetic Measurements ◽  
Steady State Kinetic ◽  
Halophilic Archaebacteria ◽  
Eukaryotic Organisms ◽  
State Kinetic

Dihydrolipoamide dehydrogenase has been discovered in the halophilic archaebacteria for the first time. The enzyme from both classical and alkaliphilic halobacteria has been investigated. (1) The enzyme specifically catalysed the stoichiometric oxidation of dihydrolipoamide by NAD+. Enzymic activity was optimal at 2 M-NaCl and was remarkably resistant to thermal denaturation. (2) The relative molecular masses (Mr) of the native enzyme from the various species of halobacteria were determined to be within the range 112000-120000. (3) The enzyme exhibited a hyperbolic dependence of catalytic activity on both dihydrolipoamide and NAD+ concentrations. From these steady-state kinetic measurements the dissociation constant (Ks) of dihydrolipoamide was determined to be 57 (+/- 5) microM. (4) The enzyme was only susceptible to inactivation by iodoacetic acid in the presence of its reducing ligands, dihydrolipoamide or NADH. The rate of inactivation followed a hyperbolic dependence on the concentration of dihydrolipoamide, from which the Ks of this substrate was calculated to be 55 (+/- 7) microM. Together with the steady-state kinetic data, the pattern of inactivations is consistent with the involvement in catalysis of a reversibly reducible disulphide bond, as has been found in dihydrolipoamide dehydrogenase from non-archaebacterial species. In eubacterial and eukaryotic organisms, dihydrolipoamide dehydrogenase functions in the 2-oxo acid dehydrogenase complexes. These multienzyme systems have not been detected in the archaebacteria, and, in the context of this apparent absence, the possible function and evolutionary significance of archaebacterial dihydrolipoamide dehydrogenase are discussed.

scholarly journals Thermodynamic and steady-state-kinetic investigation of the effect of NN′-dicyclohexylcarbodi-imide on H+ translocation by the mitochondrial cytochrome bc1 complex

1985 ◽  
Vol 225 (2) ◽  
pp. 407-411 ◽  
Author(s):  
M D Brand ◽  
M K Al-Shawi ◽  
G C Brown ◽  
B D Price
Keyword(s):  
Steady State ◽  
Thermodynamic Assessment ◽  
Cytochrome Bc1 ◽  
Kinetic Investigation ◽  
Bc1 Complex ◽  
Cytochrome Bc1 Complex ◽  
Kinetic Measurements ◽  
Steady State Kinetic ◽  
Force Ratio ◽  
State Kinetic

Steady-state kinetic measurements showed that NN′-dicyclohexylcarbodi-imide decreased the observed H+/2e ratio of H+ transport by mitochondria respiring on succinate, acting mainly at the cytochrome bc1 complex. Thermodynamic assessment of the H+/2e ratio by measuring the force ratio across the bc1 complex showed that the inhibitor did not affect H+ translocation. Possible explanations of this disagreement between methods are examined; we conclude that the inhibitor does not alter the mechanistic stoichiometry of H+ pumping by the bc1 complex.

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