Membrane ATPase from the aceticlastic methanogen Methanothrix thermophila.
A new isolate of the aceticlastic methanogen Methanothrix thermophila utilizes only acetate as the sole carbon and energy source for methanogenesis (Y. Kamagata and E. Mikami, Int. J. Syst. Bacteriol. 41:191-196, 1991). ATPase activity in its membrane was found, and ATP hydrolysis activity in the pH range of 5.5 to 8.0 in the presence of Mg2+ was observed. It had maximum activity at around 70 degrees C and was specifically stimulated up to sixfold by 50 mM NaHSO3. The proton ATPase inhibitor N,N'-dicyclohexylcarbodiimide inhibited the membrane ATPase activity, but azide, a potent inhibitor of F0F1 ATPase (H(+)-translocating ATPase of oxidative phosphorylation), did not. Since the enzyme was tightly bound to the membranes and could not be solubilized with dilute buffer containing EDTA, the nonionic detergent nonanoyl-N-methylglucamide (0.5%) was used to solubilize it from the membranes. The purified ATPase complex in the presence of the detergent was also sensitive to N,N'-dicyclohexylcarbodiimide, and other properties were almost the same as those in the membrane-associated form. The purified enzyme revealed at least five kinds of subunits on a sodium dodecyl sulfate-polyacrylamide gel, and their molecular masses were estimated to be 67, 52, 37, 28, and 22 kDa, respectively. The N-terminal amino acid sequences of the 67- and 52-kDa subunits had much higher similarity with those of the 64 (alpha)- and 50 (beta)-kDa subunits of the Methanosarcina barkeri ATPase and were also similar to those of the corresponding subunits of other archaeal ATPases. The alpha beta complex of the M. barkeri ATPase has ATP-hydrolyzing activity, suggesting that a catalytic part of the Methanothrix ATPase contains at least the 67- and 52-kDa subunits.
Inatomi
K
K
Central Research Laboratory, Mitsubishi Electric Corporation, Hyogo, Japan.
Kamagata
Y
Y
Nakamura
K
K
D000085:Acetates / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000276:immunology / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D003429:Cross Reactions; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D019605:Euryarchaeota / Q000201:enzymology*; D008565:Membrane Proteins / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D008970:Molecular Weight
D000085:Acetates; D008565:Membrane Proteins; D004024:Dicyclohexylcarbodiimide; D000251:Adenosine Triphosphatases