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Leishmania Plasma Membrane Mg21-ATPase Is a H1/K1-Antiporter Involved in Glucose Symport

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Title Leishmania Plasma Membrane Mg21-ATPase Is a H1/K1-Antiporter
Involved in Glucose Symport
 
Creator Mukherjee, Tanmoy
Mandal, Debjani
Bhaduri, Amar
 
Subject Chemistry
 
Description Experiments from other laboratories conducted with
Leishmania donovani promastigote cells had earlier indicated
that the plasma membrane Mg21-ATPase of the
parasite is an extrusion pump for H1. Taking advantage
of the pellicular microtubular structure of the plasma
membrane of the organism, we report procedures for
obtaining sealed ghost and sealed everted vesicle of defined
polarity. Rapid influx of H1 into everted vesicles
was found to be dependent on the simultaneous presence
of ATP (1 mM) and Mg21 (1 mM). Excellent correspondence
between rate of H1 entry and the enzyme
activity clearly demonstrated the Mg21-ATPase to be a
true H1 pump. H1 entry into everted vesicle was
strongly inhibited by SCH28080 (IC50 5 ;40 mM) and by
omeprazole (IC50 5 ;50 mM), both of which are characteristic
inhibitors of mammalian gastric H1,K1-ATPase.
H1 influx was completely insensitive to ouabain (250
mM), the typical inhibitor of Na1,K1-ATPase. Mg21-
ATPase activity could be partially stimulated with K1
(20 mM) that was inhibitable (>85%) with SCH28080 (50
mM). ATP-dependent rapid efflux of 86Rb1 from preloaded
vesicles was completely inhibited by preincubation
with omeprazole (150 mM) and by 5,5*-dithiobis-(2-
nitrobenzoic acid) (1 mM), an inhibitor of the enzyme.
Assuming Rb1 to be a true surrogate for K1, an ATP-dependent,
electroneutral stoichiometric exchange of H1
and K1 (1:1) was established. Rapid and 10-fold active
accumulation of [U-14C]2-deoxyglucose in sealed ghosts
could be observed when an artificial pH gradient (interior
alkaline) was imposed. Rapid efflux of [U-14C]D-glucose
from preloaded everted vesicles could also be initiated
by activating the enzyme, with ATP. Taken
together, the plasma membrane Mg21-ATPase has been
identified as an electroneutral H1/K1 antiporter with
some properties reminiscent of the gastric H1,K1-
ATPase. This enzyme is possibly involved in active accumulation of glucose via a H1-glucose symport system
and in K1 accumulation.
 
Date 2001
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/896/1/JOURNAL_OF_BIOLOGICAL_CHEMISTRY%2C__276(_8)%2C__5563%2C2001[56].pdf
Mukherjee, Tanmoy and Mandal, Debjani and Bhaduri, Amar (2001) Leishmania Plasma Membrane Mg21-ATPase Is a H1/K1-Antiporter Involved in Glucose Symport. The Journal of Biological Chemistry, 276 (8). pp. 5563-5569.
 
Relation http://dx.doi.org/10.1074/jbc.M008469200
http://www.eprints.iicb.res.in/896/