ABBOTTABAD UNIVERSITY OF SCIENCE AND TECHNOLOGY

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(Placeholder1)
Plant Physiology-I
Electrogenic Pumps-ATPases
By azan khan
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Introduction
Electrogenic pumps are primary active transporters that hydrolyze ATP and use the energy
released from ATP hydrolysis to transport ions across biological membranes leading to the
translocation of net charge across the membrane.
1.1 For example
The Na+/K+ ATPase (sodium pump) is an electrogenic pump because during every transport
cycle, it transports 3 Na+ ions out of the cell and 2 K+ ions into the cell. This leads to the
movement of one net positive charge out of the cell making this process electrogenic.
The electrical properties of plant cell membranes are quite diverse, reflecting the wide range of
environmental conditions to which plant cells are exposed. However, it appears that electrogenic
pumps almost always make important contributions to the magnitude of the membrane potential
and, in some cases, the membrane conductance.
1.2 Discussion
Early studies of membrane potentials using intracellular electrodes were summarized by
Blinks (1949). Due to technical limitations, the early work involved the use of large
freshwater or marine algal cells such as Valonia, and it was not until the development of
modern microelectrode techniques and electronics that the methods became more reliable,
beginning in the late 1950s. Initial approaches to the problem of accounting for the
magnitudes of the membrane potential and conductance and their response to changes in
external ion concentrations were influenced strongly by the success of animal physiologists
in making the simplifying assumption that the gradients of ion concentrations across the
plasma membrane were established by neutral ion pumps, i.e. pumps that would not generate
a current. It was also assumed that the electrical properties of the membrane were determined

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entirely by passive diffusion of the ions across the membrane down the gradient for each ion
that resulted from its concentration gradient and the common electrical potential gradient,
summed as the electrochemical potential gradient. The assumption of a constant electric
potential gradient (electric field) within the membrane, together with a constant partition
coefficient for an ion at both surfaces of the membrane, makes it possible to integrate the
diffusion equations across the membrane and yields an equation for the net passive flux, J.
The flux, JK, for potassium, as an example, is:
𝑗 𝑘 = −𝑃𝑘 (
𝐹𝐸
𝑅𝑇
)
[( 𝐾°+)−[ 𝐾𝑖
+] 𝑒𝑥𝑝( 𝐹𝐸
𝑅𝑇⁄ )]
{1−𝑒𝑥𝑝( 𝐹𝐸
𝑅𝑇⁄ )}
(1)
where PK is the permeability coefficient, E is the difference in electrical potential across the
membrane (the membrane potential), [Ko +] and [Ki +] are the external and internal
potassium concentrations, respectively, F is the faraday and R and T have their usual
meanings.
1.3 Further Assumption
The further assumption of a steady state, or specifically that there is no net charge accumulation
in the cell, means that the net currents carried by the individual ions must sum to zero. It leads to
the equation, usually referred to as the Goldman–Hodgkin–Katz (or GHK) equation (Goldman
1943, Hodgkin and Katz 1949), which, assuming that sodium, potassium and chloride are the
only ions transported, has the form
E= (RT/F) In
𝑃 𝑘 [ 𝐾°+]+ 𝑃 𝑁𝑎 [ 𝑁𝑎°+] + 𝑃 𝐶𝑙 [ 𝐶𝑙 𝑖
− ]
𝑃 𝑘 [ 𝐾°+]+𝑃 𝑁𝑎[ 𝑁𝑎 𝑖
+]+𝑃 𝐶𝑙[ 𝐶𝑙°
−]
(2 )
It is also possible to find an expression for the conductance in terms of the concentrations and
permeability coefficients by first applying the condition:
∑ 𝑍𝑗 𝐹𝐽𝑗 = I, (3)

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where I is the applied current, 𝑍𝑗 is the valence of the ion j, ∑ 𝑍𝑗 𝐹𝐽𝑗 and is the sum of the
currents due to the fluxes of the individual ions. Differentiating, to obtain dI/dE, gives the
conductance:
𝐺 𝑚 =[
𝐹3 𝐸
( 𝑅𝑇)2
] [
𝑤𝑦
𝑤−𝑦
] (4)
In the limit I→0, where 𝐺 𝑚 (S.𝑚2
) is the specific conductance of the membrane,
w = 𝑃𝑘 [ 𝐾°+] + 𝑃 𝑁𝑎 [ 𝑁𝑎°+] + 𝑃 𝐶𝑙 [𝐶𝑙𝑖
−
] [Nao +] and y = 𝑃 𝑘 [ 𝐾°+] + 𝑃 𝑁𝑎[ 𝑁𝑎𝑖
+] +
𝑃 𝐶𝑙[ 𝐶𝑙°
_
] . This expression was also derived by Hodgkin and Katz
Equations 1, 2 and 4 provide the basis for determining whether the electrical properties of the
membrane can be accounted for simply by passive diffusion of the ions. The permeability
coefficients are defined by Eq. (1), or partial equations for the unidirectional fluxes derived from
it. Since the membrane potential appears in all the equations, they are not independent. However,
if the assumption of passive ion diffusion is valid, the equations should give consistent results.
They can, of course, be extended to include other ions that have significant fluxes across the
membrane. The results of early attempts to apply this approach to plant cells were presented
clearly by Dainty (1962). Even at that stage, problems were becoming evident. However, the
approach does provide a sound quantitative theoretical basis on which to proceed. Thus,
permeability coefficients could be calculated from the passive components of the major ion
fluxes and used with concentration data to make predictions about the magnitude of the
membrane potential and conductance or the response to changes in external ion concentrations.
In text
Volkov, A. G. (2012). Plant Electrophysiology. Berlin: Springer Berlin.

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2..Mechanism of action
Crystal structures of the Na+
/K+
--ATPase show that ions are bound and occluded deep within
the protein about 60% of the way through the pore from the extracellular medium. Ions reach
their binding sites through hydrophilic paths called access channels. Most of the Na+
/K+
-
ATPase’s voltage dependence originates as ions move along these access channels sensing the
electric field across the membrane. The kinetic, thermodynamic and electrical properties of these
access channels and their associated occlusion/deocclusion transitions for Na+
ions have been
well characterized by both electrophysiological and spectroscopic approaches . These Na+-
dependent signals are large and relatively slow. Transient electrical signals mediated by the
binding/release of external Na+ have been particularly useful in dissecting multiple
occlusion/deocclusion events as Na+ ions are released to the extracellular medium8. The
electrogenicity of K+ binding has also been established. Nonetheless, direct measurements of
transient electrical signals mediated by K+ ions have been difficult to detect , presumably
because these transient electrical signals are small and fast. There are no available
electrophysiological recordings of K+-mediated currents associated with external
K+ binding/occlusion. However, to slow down the kinetics of these transitions, Peluffo and
Berlin substituted external K+ by Tl+, a congener K+ ion capable of being transported by the
Na+/K+-ATPases . The properties of these transient currents were consistent with ions traversing
the electric field through access channels. Further support of external K+ ions moving through
access channels comes from observations that quaternary ammonium ions, although able to
compete with external K+ binding, they are not occluded, and that the binding and unbinding
kinetics were voltage dependent.

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Using high-speed voltage clamp and large squid giant axons (>1 mm diameter), we were able to
characterize transient currents mediated by the binding and occlusion of external K+. Indeed, the
electrical signals reported here are smaller (∼5 times) and much faster (∼10 times) than the
corresponding transient currents carried by external Na+. The amount of charge moved and the
kinetics of these K+-mediated transient currents are best described by an access channel model in
which two K+ bind sequentially but they occlude simultaneously by a voltage insensitive step.
Using molecular dynamic simulations, we dissect the electrical contributions of each K+ as they
travel through the access channel to their binding sites, providing a consistent molecular picture
of the functional data.
2.1 Extracellular K+-mediated charge movement
To study the properties of external K+ binding and the associated kinetics of K+ occlusion and
deocclusion, we measured charge relaxation mediated by the Na+/K+-ATPase in squid giant
axons from Dosidicus gigas. Axons were voltage clamped with time constants of and internally
dialysed with solutions intended to restrict the Na+/K+-ATPase’s transport cycle to partial
reactions involving extracellular K+. With 1 mM extracellular K+, total membrane currents in
response to a 2-ms voltage step to −160 mV from a holding potential of 0 mV, acquired at
500 kHz, filtered at 100 kHz and recorded at equal time intervals. Na+/K+ pump-mediated
currents were extracted as the membrane current sensitive to dihydrodigitoxigenin (H2DTG), a
specific and reversible squid Na+/K+ pump inhibitor. On sudden changes in voltage to −160 mV
and back to 0 mV, H2DTG-sensitive transient currents consist of two components. Fast
(comparable to the voltage speed of the clamp; ) and slow. The latter relaxed monoexponentially
to near-zero current values (fits indicated by red solid lines) as expected by the ionic composition
of the intracellular and extracellular solutions, which prevents completion of the transport cycle.

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.
Fig. 2 Albers–Post model for the Na+/K+pump transport mechanism.

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Fig 2.1 Charge movement mediated by extracellular K+.

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iIn contrast to Na+-occlusion process where the three charge translocation components’ quantities
and time courses are tightly correlated. The fast component of K+-mediated charge movement
appears to be unrelated to K+ transport. shows four superimposed H2DTG-sensitive transient
currents in response to voltage steps from 0 to −160 mV of different durations before returning
back to 0 mV. Irrespective of the length of the voltage step, the fast pump-mediated charge
movement at the end of the pulse has similar magnitudes , suggesting that the fast component of
the charge movement does not represent a K+ binding/occlusion transition. A parsimonious
explanation of this component is a change in the electric field produced by ouabain binding In
contrast, the slow charge (Qs) increases monotonically with a time constant similar to the slow
On relaxation open circles). These results imply that the slow component of the H2DTG-sensitive
transient currents should represent the binding/occlusion of two K+.
Fig 2.1 Fast and slow components of K+ charge movement are kinetically independent.

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2.2Structural model of the Na+/K+-pump access channel for K+
. The fraction of membrane potential sensed by the binding of extracellular K+ ions can be
calculated from Δ〈QD〉, the difference in the time-averaged displacement charge of the ion-
free and ion-bound outward facing states . In the absence of a crystal structure of the outward
facing Na+/K+ pump, a model including the entire α-subunit and the transmembrane (TM)
segments of the β- and γ- subunits was generated using the crystal structures of the Ca2+ SERCA
pump as templates( Method) . In the model structure, the two ion-binding sites are accessible to
the extracellular solution via a wide aqueous channel . Site II is directly exposed to the
extracellular solution, while site I is located at the bottom of a deep binding cleft with
coordination provided by acidic side chains . The binding of the first K+ ion in the experiment
may reflect occupancy of the two sites in rapid exchange. Assuming it is equally likely that the
first K+ binds to site I or II, the calculated λ1 and λ2 are 0.49±0.12 and 0.37±0.20, respectively.
These values are also consistent with estimates based on a linear response approximation
(equation (12)), which allows one to visualize the spatial dependence of the applied membrane
potential (Methods). Remarkably, the calculations are close to the experimentally determined
values (0.46 and 0.27; Further support for the structural model is provided by comparing with a
recent crystal structure of the Na+/K+ pump E2 state partially open to the extracellular side with
bound ouabain. MD simulations of this structure with ouabain and Mg2+ removed showed
spontaneous rebinding of Na+ to the binding site, leading to the suggestion that this crystal
structure resembles the outward facing state of the pump. The model and the crystal structure
show similarity, especially in the TM region, where the backbone root mean squared deviation
is 2.7 Å. The computational transition pathway, linking the crystal structure 2ZXE (occluded
with bound K+ to the model of the outward facing P–E2 K2 state , displays the structural

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rearrangements expected to occur during the occlusion/deocclusion process. Structural changes
in the TM domain mostly involve the M1–M4 helices in the α subunit. More specifically, M1–
M3 undergoes a piston-like motion and is pulled towards the intracellular matrix. This makes
room for the extracellular portion of M4 (residues W317 to N331), which then tilts and opens up
an aqueous channel between M4 and M6 for ions reaching the binding site . The cross-sectional
radius of the water-filled path on the occlusion/deocclusion process . This path involves residues
F323, G326, A330, E334 on M4 and T804, I807, L808 and D811 along M6. Interestingly,
several of these residues are known to be along the open conducting ion channel arising in the
palytoxin-bound conformation.
Fig 2.2.Extracellular occlusion–deocclusion of the Na+/K+-ATPase in the presence of bound
𝐊+

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3. Role in transport of solutes
3.1.. Electrochemical gradients
concentration gradients, in which a substance is found in different concentrations over a region
of space or on opposite sides of a membrane. However, because atoms and molecules can form
ions and carry positive or negative electrical charges, there may also be an electrical gradient, or
difference in charge, across a plasma membrane. In fact, living cells typically have what’s called
a membrane potential, an electrical potential difference (voltage) across their cell membrane.
Fig 3. Electrochemical gradient

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An electrical potential difference exists whenever there is a net separation of charges in space. In
the case of a cell, positive and negative charges are separated by the barrier of the cell
membrane, with the inside of the cell having extra negative charges relative to the outside. The
membrane potential of a typical cell is -40 to -80 millivolts, with the minus sign meaning that
inside of the cell is more negative than the outside. The cell actively maintains this membrane
potential, and we’ll see how it forms in the section on the sodium-potassium pump.
As an example of how the membrane potential can affect ion movement, let’s look at sodium and
potassium ions. In general, the inside of a cell has a higher concentration of potassium (K+)
start
superscript, plus, end superscript) and a lower concentration of sodium (Na+)start superscript,
plus, end superscript) than the extracellular fluid around it.
a. If sodium ions are outside of a cell, they will tend to move into the cell based on both
their concentration gradient (the lower concentration of (Na+) start superscript, plus, end
superscript in the cell) and the voltage across the membrane (the more negative charge on
the inside of the membrane).
b. Because( K+))
start superscript, plus, end superscript is positive, the voltage across the
membrane will encourage its movement into the cell, but its concentration gradient will
tend to drive it out of the cell (towards the region of lower concentration). The final
concentrations of potassium on the two sides of the membrane will be a balance between
these opposing forces.
c. The combination of concentration gradient and voltage that affects an ion’s movement is
called the electrochemical gradient.

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3.2. Active transport: moving against a gradient
To move substances against a concentration or electrochemical gradient, a cell must use energy.
Active transport mechanisms do just this, expending energy (often in the form of ATP) to
maintain the right concentrations of ions and molecules in living cells. In fact, cells spend much
of the energy they harvest in metabolism to keep their active transport processes running. For
instance, most of a red blood cell’s energy is used to maintain internal sodium and potassium
levels that differ from those of the surrounding environment.
Active transport mechanisms can be divided into two categories. Primary active
transport directly uses a source of chemical energy (e.g., ATP) to move molecules across a
membrane against their gradient. Secondary active transport (cotransport), on the other hand,
uses an electrochemical gradient – generated by active transport – as an energy source to move
molecules against their gradient, and thus does not directly require a chemical source of energy
such as ATP.
3.3 Primary active transport
One of the most important pumps in animal cells is the sodium-potassium pump, which moves
Na+start superscript, plus, end superscript out of cells, and K+start superscript, plus, end
superscript into them. Because the transport process uses ATP as an energy source, it is
considered an example of primary active transport.

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Not only does the sodium-potassium pump maintain correct concentrations of Na+start
superscript, plus, end superscript and K+start superscript, plus, end superscript in living cells, but
it also plays a major role in generating the voltage across the cell membrane in animal cells.
Pumps like this, which are involved in the establishment and maintenance of membrane voltages,
are known as electrogenic pumps. The primary electrogenic pump in plants is one that pumps
hydrogen ions (H+start superscript, plus, end superscript) rather than sodium and potassium.
3.4 The sodium-potassium pump cycle
Fig 3.4 the sodium_ potassium pump cycle
The sodium-potassium pump transports sodium out of and potassium into the cell in a repeating
cycle of conformational (shape) changes. In each cycle, three sodium ions exit the cell, while two
potassium ions enter. This process takes place in the following steps:
1. To begin, the pump is open to the inside of the cell. In this form, the pump really likes to bind
(has a high affinity for) sodium ions, and will take up three of them.

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2. When the sodium ions bind, they trigger the pump to hydrolyze (break down) ATP. One
phosphate group from ATP is attached to the pump, which is then said to be phosphorylated.
ADP is released as a by-product.
3. Phosphorylation makes the pump change shape, re-orienting itself so it opens towards the
extracellular space. In this conformation, the pump no longer likes to bind to sodium ions (has a
low affinity for them), so the three sodium ions are released outside the cell.
4. In its outward-facing form, the pump switches allegiances and now really likes to bind to (has a
high affinity for) potassium ions. It will bind two of them, and this triggers removal of the
phosphate group attached to the pump in step 2.
5. With the phosphate group gone, the pump will change back to its original form, opening towards
the interior of the cell.
6. In its inward-facing shape, the pump loses its interest in (has a low affinity for) potassium ions,
so the two potassium ions will be released into the cytoplasm. The pump is now back to where it
was in step 1, and the cycle can begin again.
3.5 Secondary active transport
The electrochemical gradients set up by primary active transport store energy, which can be
released as the ions move back down their gradients. Secondary active transport uses the energy
stored in these gradients to move other substances against their own gradients.
As an example, let's suppose we have a high concentration of sodium ions in the extracellular
space (thanks to the hard work of the sodium-potassium pump). If a route such as a channel or

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carrier protein is open, sodium ions will move down their concentration gradient and return to
the interior of the cell.
In secondary active transport, the movement of the sodium ions down their gradient is coupled to
the uphill transport of other substances by a shared carrier protein (a cotransporter). For
instance, in the figure below, a carrier protein lets sodium ions move down their gradient, but
simultaneously brings a glucose molecule up its gradient and into the cell. The carrier protein
uses the energy of the sodium gradient to drive the transport of glucose molecules.

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Fig 3.5 secondary active transport
In text
Friedman, M. H. (2008). Active Transport. Principles and Models of Biological Transport,
1-39. doi:10.1007/978-0-387-79240-8_5
4 . ATPase-proton pumps of plasma membrane and vacuolar membranes
( tonoplast)
One of the major active ion transport systems in plant plasma membrane is the electrogenic
H+ ATPase (PM-type H+ pump). The PM-type H+ pump generates an electrochemical gradient
for H+ across the plasma membrane and enables the cell to transport various kinds of nutrients,
amino acids, sugars and other materials from the extracellular spaces. Structure and regulatory
mechanism of the PM-type H+ pump have been revealed by recent molecular studies. It is

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demonstrated that the PM-type H+ pump has the autoinhibitory domain at the C-terminal region
The activity of the H+ pump is suggested to be regulated by phosphorylation or
dephosphorylation of the C-terminal region and the binding with the 14-3-3 protein.
4.1 𝐇+
_Pump Activation
Patch-clamp studies allow investigation of pump dynamics by direct measurement of the
electrogenic current (3, 50, 167). Furthermore, in whole-cell recordings the requirement of
cytoplasmic substrates for pump activation can be studied effectively. Therefore, patch-clamp
studies appear to be ideal for the investigation of the modulation of H+ pumps . New insights
into pump activation by blue light and red light and by the hormone-like compound fusicoccin
have been obtained. In guard cells, low fluence rates of blue light and high fluence rates of red
light can activate plasma membrane H+ pumps by different mechanisms . Blue-light activation
of pumps follows the initial exposure to light by a marked delay of approximately 30 sec . In
contrast, exposure to red light activates H+ pumps without a measurable delay . Both
mechanisms require cytoplasmic substrates other than Mg-ATP for maximal pump activity . By
using nonperfused guard cells to prevent loss of cytoplasmic factors it was found that blue light-
activated pump currents are increased 5-10 times by yet unidentified cytoplasmic substrates .
Moreover, the magnitude of blue light-activated pump currents measured in nonperfused guard
cells was found sufficient to activate IK+,in channels and drive K+ uptake required for stomatal
opening . The current-voltage relationship of H+ pumps in guard cells was calculated under the
assumption that the H+ pump inhibitor cyanide has no effect on channel conductances .
However, more recent data suggest that cyanid also blocks K+ channels. Biochemical studies
provide evidence that activation of H+ pumps by auxin and photonastic light stimuli may involve

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the pathway of phosphoinositol lipid metabolism . Furthermore, a cytoplasmic acidification that
preceded membrane hyperpolarization was observed in response to auxin.
Proton transport across biological membranes is a central process in many energy conversion
reactions in the cell. In the vacuolar membrane protons are translocated by electrogenic ATPases
and pyrophosphatases (PPj-ases). The two phosphatases in the vacuolar membrane are physically
distinct enzymes as confirmed by chromatographic separation and by their ionic specificitiese.g.
activation by halides or cations.
With the development of techniques for the preparation of sealed membrane vesicles from
various plant tissues in combination with pH- and voltage-sensing probes, several laboratories
have demonstrated H+ translocating phosphatases associated with fractions of the vacuolar
membrane.
(b) directly measure potentials and currents generated by the proton pumps; and (c) determine
the stoichiometry and thermodynamics of the pump reactions.
4.2 H+ ATPases
The "vacuolar type ATPase" is the most widespread H+ ATPase in eukaryotic cells . It is present
in lysosomes , vacuoles of plants and fungi, clathrin-coated vesicles, synaptic vesicles, and
several secretory granules. All H+ ATPases belonging to this group bear structural homologies
and appear to be composed of three major polypeptides . Among them the 69-kO polypeptide
revealed the highest degree of homology to the H+ ATPase of archae bacteria.
A common characteristic of these enzymes is that they catalyze the same overall reaction,
nH+ cytopas,m + ATP --> nH+ vacuole + ADP + Pi
and utilize the free energy of this reaction for the transport of H+ against an electrochemical
gradient. The factor n signifies the number of Hi- translocated by the enzyme per one A TP or

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PPj hydrolyzed. For the H+ ATPase Bennett & Spanswick deduced an ATP/H+ ratio of 2 from
kinetic measurements on A TP-hydrolysis and pH change using vacuole vesicles. A more
detailed analysis of the pump stoichiometries of both the ATPase and PPj-ase can be obtained by
measuring the reversal potential of the pump currents. Under given phosphate potentials in the
"cytoplasmic solution" the direction of the proton current through the pump can be recorded as a
function of A pH increase across the vacuolar membrane. Upon the application of Mg-ATP to
the cytoplasmic face of a wholevacuole, a rapid depolarization of 30-70 m can be observed,
indicating the presence of an inwardly directed electrogenic ATPase . This membrane potential is
immediately abolished by the H+ -ATPase inhibitor tributyltin or by the protonophore
carbonylcyanid-dichlorphenylhydrazon Flower panel aindicating that the ATP-induced shift in
membrane potential was generated by H+ currents into the vacuole. When the ATP concentration
was increased stepwise, the pump current also increased stepwise, reaching saturation at 5-10
mM Mg-ATP. Km values with respect to ATP were about 0.6 mM for Beta vulgaris taproot.
Under physiological conditions, such as 1 mM ATP in the cytoplasm and a pH-gradient of 2-3
pH units across the vacuolar membrane , the H+ -ATPase will produce a H+ current of about 50
pA/vacuole ( 1-5 /LA/cm2; 24, 49, 50). Assuming that anion fluxes shortcircuit the H+ current,
vacuoles (e.g. of barley mesophyll) can accumulate 0. 1f mol malate2- S-I at the expense of 2
protons per malate--comparable to values found for CAM plants . This estimate demonstrates
that the ATPase provides enough current to drive malate accumulation in the vacuole during
photosynthesis . The H+ -ATPase of the sugar beet taproot is able to pump against a 1Q4-fold
H+ -gradient at a membrane potential of 0 mV (pH 7.8 at the cytoplasmic side and pH 3.8 on the
vacuolar side. These experiments indicate that the proton binding sites at the cytoplasmic mouth
of the proton channel of the ATPase complex must possess a PK above 7.8 and that the PK of

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H+ release must have been below 3.8 at its vacuolar face. A proton binding site of PK < 3.8 may
indicate the presence of carboxylates , because only these groups have a PK low enough to
release protons at pH values < 4, a task that the vacuolar H+-ATPase in the lemon fruit can
certainly achieve .
Fig 4.2. H+ ATPases

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5.Types of ATPases
The four classes of ATP-powered transport proteins. P-class pumps are composed of two
different polypeptides, α and β, and become phosphorylated as part of the transport cycle. The
sequence around the phosphorylated residue, located in in the larger α subunits, is homologous
among different pumps. F-class and V-class pumps do not form phosphoprotein intermediates.
Their structures are similar and contain similar proteins, but none of their subunits are related to
those of P-class pumps. All members of the large ABC superfamily of proteins contain four core
domains: two transmembrane (T) domains and two cytosolic ATP-binding (A) domains that
couple ATP hydrolysis to solute movement. These core domains are present as separate subunits
in some ABC proteins , but are fused into a single polypeptide in other ABC proteins.
Fig 5. Types of ATPases

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1. P-class ionpumps
P-class ion pumps contain a transmembrane catalytic α subunit, which contains an ATP-binding
site, and usually a smaller β subunit, which may have regulatory functions. Many of these pumps
are tetramers composed of two α and two β subunits. During the transport process, at least one of
the α subunits is phosphorylated and the transported ions are thought to move through the
phosphorylated subunit. This class includes the Na+/K+ ATPase in the plasma membrane, which
maintains the Na+ and K+ gradients typical of animal cells, and several Ca2+ ATPases,
which pump Ca2+ ions out of the cytosol into the external medium or into the lumen of
the sarcoplasmic reticulum (SR) of muscle cells. Another member of the P class, found in acid-
secreting cells of the mammalian stomach, transports protons (H+ ions) out of and K+ ions into
the cell. The H+ pump that maintains the membrane electric potential in plant, fungal, and
bacterial cells also belongs to this class.
2. F-class and V-class ionpumps
F-class and V-class ion pumps are similar to each other but unrelated to and more
complicated than P-class pumps. F- and V-class pumps contain at least three kinds of
transmembrane proteins and five kinds of extrinsic polypeptides that form the
cytosolic domain. Several of the transmembrane and extrinsic subunits in F-class and V-class
pumps exhibit sequence homology, and each pair of homologous subunits is thought to have
evolved from a common polypeptide.
All known V and F pumps transport only protons in a process that does not involve a
phosphoprotein intermediate. V-class pumps generally function to maintain the low pH of
plant vacuoles and of lysosomes and other acidic vesicles in animal cells by using the energy

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released by ATP hydrolysis to pump protons from the cytosolic to the exoplasmic face of
the membrane against the proton electrochemical gradient. F-class pumps are found in
bacterial plasma membranes and in mitochondria and chloroplasts. In contrast to V pumps,
they generally function to power the synthesis of ATP from ADP and Pi by movement of
protons from the exoplasmic to the cytosolic face of the membrane down the proton
electrochemical gradient. Because of their importance in ATP synthesis in chloroplasts and
mitochondria.
3. ABC (ATP-binding cassette) superfamily
The final class of ATP-powered transport proteins is larger and more diverse than the other
classes. Referred to as the ABC (ATP-binding cassette) superfamily, this class includes more
than 100 different transport proteins found in organisms ranging from bacteria to humans.
Each ABC protein is specific for a single substrate or group of related substrates including
ions, sugars, peptides, polysaccharides, and even proteins. All ABC transport proteins share a
common organization consisting of four “core” domains: two transmembrane (T) domains,
forming the passageway through which transported molecules cross the membrane, and two
cytosolic ATP-binding (A) domains. In some ABC proteins, the core domains are present in
four separate polypeptides; in others, the core domains are fused into one or two multidomain
polypeptides.
All classes of ATP-powered pumps have one or more binding sites for ATP, and these are
always on the cytosolic face of the membrane . Although these proteins are often called
ATPases, they normally do not hydrolyze ATP into ADP and Pi unless ions or other molecules
are simultaneously transported. Because of the tight coupling between ATP hydrolysis and
transport, the energy stored in the phosphoanhydride bond is not dissipated. Thus ATP-powered

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transport proteins are able to collect the free energy released during ATP hydrolysis and use it to
move ions or other molecules uphill against a potential or concentration gradient.
The energy expended by cells to maintain the concentration gradients of Na+, K+, H+, and
Ca2+ across the plasma and intracellular membranes is considerable. In nerve and kidney cells,
for example, up to 25 percent of the ATP produced by the cell is used for ion transport; in human
erythrocytes, up to 50 percent of the available ATP is used for this purpose. In cells treated with
poisons that inhibit the aerobic production of ATP (e.g., 2,4-dinitrophenol), the ion concentration
inside the cell gradually approaches that of the exterior environment as the ions move
through plasma membrane channels down their electric and concentration gradients. Eventually
treated cells die: partly because protein synthesis requires a high concentration of K+ ions and
partly because in the absence of a Na+ gradient across the cell membrane, a cell cannot import
certain nutrients such as amino acids. Studies on the effects of such poisons provided early
evidence for the existence of ion pumps. In this section, we discuss in some detail examples of
the P, V, and ABC classes of ATP-powered pumps.
In text
Lodish, H. F. (2016). Molecular cell biology. New York, NY: Freeman.

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Refrences
Tsutsui, I., & Ohkawa, T. (2001). Regulation of the H+ Pump Activity in the Plasma
Membrane of Internally Perfused Chara corallina. Plant and Cell Physiology, 42(5),
531-537. doi:10.1093/pcp/pce068

28. 27

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