Inhibition of ATPase

Apart from the action upon Na+ channels, p,p -DDT and its metabolites can have certain other toxic effects. It has been reported that p,p -DDT can inhibit certain ATPases (see EHC 83). In fish, the inhibition of ATPases can affect osmoregulation. 

During digestion at 35°C in the presence of Ca the formation of the Aja and Aib fragments was accompanied by loss of ATPase activity, but significant E P formation was preserved in the A a and Aib fragments [235]. The E P formed in the A b + B complex was ADP sensitive (EiP), even in the absence of KCl, suggesting that its conversion into the ADP-insensitive E2P was impaired. This effect may account for the inhibition of ATPase activity after cleavage of the T3a and Tab sites [123]. 

Some of the antibodies directed against the B fragment of the Ca -ATPase caused moderate inhibition of ATPase activity and Ca transport the inhibition usually did not exceed 50%, even at high antibody Ca -ATPase ratios where the antigenic sites are expected to be fully saturated [285,302,304],... 

The powerful inhibition of Ca " transport without inhibition of ATPase activity seen with polyclonal anti-ATPase sera in earlier studies [305] was probably due to complement dependent lesion of the membrane that permitted the leakage of accumulated calcium [306-308], The scarcity of inhibitory antibodies may imply that the active site of the Ca - ATPase is either inaccessible to antibodies or poorly antigenic, perhaps due to a unique secondary structure. 

Reaction of purified Ca " -ATPase with 0.3 mM NBD-Cl in the presence of 1 mM AMP-PNP and 1 mM CaCl2 caused inhibition of ATPase activity with the incorporation of 2= 15 nmol NBD-Cl per mg protein [335]. The inhibition was attributed to the binding of 7-8 nmol NBD-Cl/mg enzyme protein, corresponding to = 1 mol NBD-Cl per mol ATPase. The NBD-labeled enzyme was digested with pepsin and several NBD-labeled peptides were isolated [335]. All peptides contained the Gly-X (Cys) sequence that occurs only in one place in the Ca -ATPase, i.e., at Gly343-Cys344. Therefore NBD-Cl reacts with the same cysteine 344 residue that is also modified by maleimide derivatives [319]. The NBD modified enzyme had only 5-10% of the ATPase activity of the control ATPase, but the steady state concentration of the phosphoenzyme intermediate was only slightly reduced [335]. The Ca ... 

Chemical modification studies with fluorescein-5 -isothiocyanate support the proximity of Lys515 to the ATP binding site [98,113-117,212,339]. Fluorescein-5 -isothiocyanate stoichiometrically reacts with the Ca -ATPase in intact or solubilized sarcoplasmic reticulum at a mildly alkaline pH, causing inhibition of ATPase activity, ATP-dependent Ca transport, and the phosphorylation of the Ca " -ATPase by ATP the Ca uptake energized by acetylphosphate, carbamylphos-phate or j -nitrophenyl phosphate is only partially inhibited [113,114,212,339]. The reaction of -ATPase with FITC is competitively inhibited by ATP, AMPPNP, TNP-ATP, and less effectively by ADP or ITP the concentrations of the various nucleotides required for protection are consistent with their affinities for the ATP binding site of the Ca -ATPase [114,212,340]. 

Similarly, the rate of inhibition of phosphoenzyme formation by diethylpyrocarbonate (DEPC) was much slower than the loss of ATPase activity [368], Even when the reaction approached completion with more than 90% inhibition of ATP hydrolysis, about 70% of the Ca -ATPase could still be phosphorylated by ATP (2.3nmoles of E P/mg protein). The remaining 30% of E P formation and the corresponding ATPase activity was not reactivated by hydroxylamine treatment, suggesting some side reaction with other amino acids, presumably lysine. When the reaction of the DEPC-modified ATPase with P-ATP was quenched by histidine buffer (pH 7.8) the P-phosphoenzyme was found to be exceptionally stable under the same conditions where the phosphoenzyme formed by the native ATPase underwent rapid hydrolysis [368]. The nearly normal phosphorylation of the DEPC-trea-ted enzyme by P-ATP implies that the ATP binding site is not affected by the modification, and the inhibition of ATPase activity is due to inhibition of the hydrolysis of the phosphoenzyme intermediate [368]. This is in contrast to an earlier report by Tenu et al. [367], that attributed the inhibition of ATPase activity by... 

NCD-4 is a nonfluorescent carbodiimide derivative that forms a fluorescent adduct with the Ca -ATPase, accompanied by inhibition of ATPase activity and phos-phoenzyme formation [376-378]. Ca protected the enzyme against the inhibition by NCD-4 and reduced the extent of labeling, suggesting that the reaction may involve the Ca " " binding site. The stoichiometry of the Ca -protected labeling was i 2mole/mol ATPase. The fluorescence emission of the modified Ca -ATPase is consistent with the formation of a protein bound A-acylurea adduct in a relatively hydrophobic environment. After tryptic proteolysis of the NCD-4 labeled ATPase the fluorescence was associated with the A2 band of 24 kDa [376,379]. 

The influence of these phenolic acids on electrical potentials may reflect effects on either the diffusion potential or the electrogenic potential of plant root cells. Influence on the electrogenic component could result from inhibition of ATPases which generate the electrogenic component or from reductions in the substrate (ATP) for the ATPases. 

AD-9161 represents the most potent of a novel but mechanistically similar class of irreversible H /K -ATPase inhibitors (134). Compounds of this type were designed based on the finding that the oxoisothiazolo[5,4-6]pyridine (3) displayed potent inhibition of ATPase in vitro but failed to achieve inhibition of gastricacid secretion in vivo (135). This disparity was ascribed to the rapid and preferential attachment to and inactivation of (3), by thiol groups located outside the region of the ATPase enzyme in the parietal cells. AD-9161 is considerably more stable than (3)and displays potency comparable to that of omeprazole both in vitro and in vivo. 

Dibromobimane >50% inhibition of ATPase activity. Could be rescued by preincubation [196]... 

Apart from His, Cys and Ser, the hydroxyamino acid threonine can also be a residue in the active site of phosphatases the Thr214Ala mutant of Na+K+-ATPase exhibits a drastically reduced vanadate affinity for inhibition of ATPase activity,I ] suggesting that Thr is essential in vanadate binding. In the light of what has been said above, threonine very probably coordinates to vanadium into one of the axial positions in much the same manner as the other amino acid residues. 

In early 2002, we determined the crystal structure of Nt-Hsp90 from both the a- and (3-forms [19]. There was no measurable difference in the inhibition of ATPase activity by the initial tool compound (PU3, compound 2 [20]) between... 

The inhibition of ATPase with a series of ATP analogues has been studied. Among the compounds examined, adenosine-5 -bromoacetylphosphonate inhibited the enzyme at a fairly high rate, both in the presence and in the absence of ATP. ... 

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