Comparative Analysis of Mg-Dependent and Mg-Independent
Received: 4 June 2014 / Accepted: 11 September 2014 / Published online: 16 October 2014
Ó Springer Science+Business Media New York 2014
Abstract The comparative analysis between two enzymes,
Mg-dependent and Mg-independent HCO
studied in synaptosomal and microsomal membrane fractions
of albino rat brain, using the method of kinetic analysis of the
multi-sited enzyme systems. Therefore, it can be inferred that
ATPase belongs to the group of
‘‘P-type’’ transporting ATPases. Mg-independent HCO
ATPase with its kinetic properties may be attributed to the
group of ‘‘Ecto’’ ATPases.
Keywords Mg-dependent and Mg-independent HCO
ATPases Á P-type ATPase Á Ecto-ATPase Á Enzyme
kinetic Á Multi-sited enzyme system
ATPase is known as Mg-dependent enzyme system
activated by the HCO
E.C.188.8.131.52.). Mg-dependent HCO
ATPase is found in the
plasma membrane of animal and plant cells (Ivashenko 1977;
Izutsu and Siegel 1980). The participation of this enzyme in
the process of active transport of bicarbonate ion through the
membrane is supposed (Simon and Thomas 1972; Gassnez
and Komnick 1981; Tsakadze and Koshoridze 1976).
The asymmetric distribution of anions (mainly HCO
) in the membrane (out [[ in) accounts for their passive
transport down the concentration gradient whose reverse sys-
tem appears to be the existence of active transport mechanism.
Transport ATPases are known to have a particularly important
role in the cell functioning. Providing the asymmetric
arrangement of ions in the membrane at the expense of ATP
hydrolysis, they represent a complex biological machine.
But the ATPase system accomplishing ATP hydrolysis
without Mg ion (Mg
been observed in synaptosomal and microsomal membrane
fractions of the albino rat brain (Leladze et al. 2010).
There is no available information on Mg-independent
ATPase in the literature. According to our data, the
activation of ATPase reaction ﬁxed by HCO
the participation of Mg ions should not be attributed to the
so-called P-type transport ATPases system (Dzneladze
et al. 2012). Thus, the place of Mg-independent HCO
ATPase is unclear in modern classiﬁcation.
The goal of our work was to reveal the mechanism of
action of Mg-dependent and Mg-independent HCO
ATPase using the method of kinetic analysis of the multi-
sited enzyme systems (Kometiani 2007), as well as to
perform their comparative analysis and to establish a place
of each of them in ATPase classiﬁcation.
Materials and Methods
The synaptosomal and microsomal membrane fractions
obtained from albino rat brains via the method of differ-
ential centrifugation in a sucrose gradient [1.2–0.9 M
sucrose] (Kometiani et al. 1984) was an enzymatically
active material. ATPase activity (V) was assessed by the
amount of isolated inorganic phosphorus (presumed to
originate from ATP dissociation) per mg protein per hour.
Protein concentration was measured by the Lowry method
(Lowry et al. 1951). Inorganic phosphorus was measured
using a modiﬁed method of (Kazanova and Maslova 1984).
S. Dzneladze (&) Á L. Tsakadze Á M. Leladze Á E. Nozadze Á
N. Arutinova Á L. Shioshvili Á G. Chkadua
Beritashvili Center of Experimental Biomedicine, 14 Gotua str,
0160 Tbilisi, Georgia
J Membrane Biol (2015) 248:53–58