Dicyclohexylcarbodiimide-sensitive ATPase in Halobacterium saccharovorum

Halobacterial adenosine triphosphatases and the adenosine triphosphatase from Halobacterium saccharovorum

“…These rates were significantly different from those previously reported [9,10]. Contrary to what was observed with membranes from H. halobium [9], ADP was not hydrolyzed and, in fact, inhibited ATP hydrolysis in a competitive manner [15]. Since competitive inhibitions occur when compounds react with the same form of the enzyme, the inhibition by ADP indicated that ATP hydrolysis proceeded through an enzyme-ADP intermediate.…”

“…Maximum ATPase activity occurred when the concentration of NaC1 was 3.5 M or greater. At lower salt concentrations, the response of ATP hydrolysis to NaC1 concentration showed a sigmoid dependence with little if any activity detected in the presence of 1 M NaC1 [15]. This probably reflected assay conditions which simultaneously measured the rapid inactivation and low ATPase activity.…”

“…Membranes prepared from a variety of extreme halophiles exhibited little, if any, ATPase activity in the absence of detergents. Of the detergents tested (such as Triton X-100, sodium deoxycho- Table 1 The effect of Triton X-100 on the ATPase activity in membranes from H. saccharovorum Membranes were prepared by suspending early stationary phase cells in 50 mM Tris-HC1 buffer pH 8.0 that was 4 M with respect to NaC1, and disrupting them in a French pressure cell as previously described [15]. The complete reaction mixture contained the following additions in a total volume of 1 ml: 50 /~mol Tris-HC1, pH 9; 4 mmol NaCI; 5/~mol MgCI2; 10/Lmol ATP (added as the MgATP complex), 0.1% Triton X-100 (v/v); and membranes containing 0.9 mg protein.…”

“…Since competitive inhibitions occur when compounds react with the same form of the enzyme, the inhibition by ADP indicated that ATP hydrolysis proceeded through an enzyme-ADP intermediate. The membrane-bound enzyme was also inhibited by 4-chloro-7-nitrobenzofurazan [15], a reagent that inhibits FoF1-ATPases by reacting with a tyrosine residue on the fl-subunit [17]. The membrane-bound enzyme was not inhibited by phosphate (40 mM), oubain (2 mM), orthovanadate (5 mM), Dio 9 (120 #g/mg membrane protein) or azide (2 mM) [15].…”

Kinetic regulation of an alkalinep-nitrophenylphosphate phosphatase fromHalobacterium salinarumin low water system by Mn2+and monovalent cations

“…Selective effects of Mn 2+ have been reported for different halophilic enzymes [14–17]. An ATPase from Halorubrum saccharovorum showed a hysteretic behaviour, and the kinetic model proposed involves two forms of the enzyme, an initial form and a final form with distinct hydrolytic properties [16].…”