NADH oxidase from the extreme thermophile Thermus aquaticus YT-1. Purification and characterisation.
A protein with NADH oxidase activity from the extreme thermophile Thermus aquaticus YT-1 was purified and characterised. The enzyme was found to have a relative molecular mass of 110,000 and be composed of two subunits of identical size. FAD was found to be present at a concentration of 0.7 mol/mol dimer and was required for activity. During the oxidation of NADH, oxygen uptake takes place with the production of hydrogen peroxide. The enzyme had, with the exception of a higher glutamic acid and tryptophan content, a similar amino acid composition as the NADH oxidase isolated from the mesophile Bacillus megaterium. Purified NADH oxidase was found to have a Km of 39 microM for beta-NADH and a Vmax of 4.68 mumol NADH mg-1 min-1 and was still active at 95 degrees C. Enzymatic activity was found to be independent of pH between 5.0 and 10.5.
Cocco
D
D
Institute of Biological Chemistry, Faculties of Pharmacy and Medicine, University of Cagliari, Italy.
Rinaldi
A
A
Savini
I
I
Cooper
J M
JM
Bannister
J V
JV
D000596:Amino Acids / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005415:Flavins / Q000032:analysis; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009097:Multienzyme Complexes / Q000302:isolation & purification; D009247:NADH, NADPH Oxidoreductases / Q000302:isolation & purification; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000302:isolation & purification*; D013824:Thermus / Q000201:enzymology*
D000596:Amino Acids; D005415:Flavins; D009097:Multienzyme Complexes; D006861:Hydrogen Peroxide; D010544:Peroxidases; C021845:NAD+ peroxidase; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases