The Na+/K+ ATPase (NKA)

The Na+/K+-ATPase
(NKA) is a ubiquitously expressed P-type ATPase. According to the chemi-osmotic
theory (Mitchell 1961), its principal role is to maintain the ionic gradients
of sodium (Na+) and potassium (K+)
across the plasma membrane by adenosine triphosphate (ATP) hydrolysis-driven
active transport. A typical cell keeps a resting membrane potential Vm
of -70 mV. Based on the Nernst equation, K+ will tend to flow out of
the cell,  since the K+
equilibrium, or Nernst potential (E = -91 mV) is more negative than the
transmembrane potential (Suhail, 2010), whereas Na+ ions are driven
into the cell from a large extracellular pool. NKA’s P-type classification
refers to phosphorylation-dependent conformational changes it undergoes between
E1 and E2 during its canonical cycle (the Post-Albers
Cycle), E1 for the active movement of 3Na+ ions out of
the cell, and E2 for 2K+ ions into the cytosol (Post et
al., 1960). The electrochemical potential gradients created for both ion
species are essential for cellular functions such as secondary active
transport, excitability, and volume regulation (Apell 2014).  Without these functions eukaryotic life would
not be sustainable.  Sodium ions are
toxic because they take the place of potassium, which is required for the biosynthesis
of proteins.  Moreover, while sodium
specifically inhibits the normal function of certain proteins (Garcia-Salcedo,
2006), its increase has been related to transcriptional upregulation (Xie et al., 2001).  Therefore, NKA due to its ubiquitous and
essential nature, has become a prevalent target for emerging

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pharmaceuticals (Yatime et al., 2009). Their
disruption of the electrochemical gradient, as some studies have shown, lowers
mortality rates in cancer patients treated with cardiac glycosides, and particularly
in women with breast cancer (Garcia, 2015). 
NKA is also involved in hypertension, salt balance, cardiovascular and
renal disorders, sperm capacitation, cell volume regulation, apoptosis,
rheumatoid arthritis, sepsis, neurological disorders, lung edema clearance and
preeclampsia (Suhail, 2010). As NKA activity influences such a broad range of
ailments, this suggest regulatory mechanisms associated with the interaction
with cardiac glycosides.

NKA is a heterodimer
consisting of ? and ? subunits. Of these two subunits, the ? isoform, the
catalytic unit required for ion transport, exists in four isoforms (?1–?4) whereas
three ? subunit isoforms (?1–?3) have been identified thus far. One of seven ?
subunits from the family of the FXYD (phenylalanine-X-tyrosine-aspartate)
proteins (Geering, 2006) is involved in Na pump
regulation by interacting with ? and ? subunits thus forming functionally a
trimeric oligomer (Dietze, 2013).  Each
NKA ? subunit is a multi-spanning transmembrane protein responsible for its catalytic
and transport properties, as well as for providing binding sites for cations,
ATP, and cardiotonic steroids (CTS) (Pierre, 2006).  CTS’s have been in use for over two centuries
from a clinical and research perspective. 
Due to their pharmacologically known selectivity, CTS can be used to
identify how a wide variety of cells react when they target NKA and form the
NKAR-CTS complex.

Cardiotonic Steroids (Ouabain)

CTS compounds have a
generic structure which consists of a steroid core and a lactone ring considered
the pharmacophoric moiety responsible for their activity. The steroid core is
double-substituted with an unsaturated lactone ring at carbon (C) position 17
and a sugar portion at position C 3 (Prassas et al., 2008).  Figure 1 shows the differing structural features
of cardenolides vs. bufadienolides based on the 5 or 6-member lactone ring
bound to the 17-position carbon of the steroid core.