Membrane transport ATPases are among the most important consumers of ATP in cells. Ion regulation across selectively-permeable cellular membranes requires active pumping against concentration gradients, whereby transport ATPases couple ATP hydrolysis with this transfer of ions. The biochemical processes involved in metabolic depression often involve a reduction in both locomotor activity and the energy required to maintain cellular homeostasis. Ion gradients across cell membranes are crucial for survival, even in a metabolically depressed state; however, the cost involved in maintaining Na+ and K+ gradients is high and is regulated by the ATP-driven ion pumps (Na+/K+-ATPase) and ion-specific leak channels. Though the cost of maintaining ion gradients is unquestionably high, mechanisms such as channel arrest and concurrent downregulation of ATPases could lower this cost to a manageable level. The theory of channel arrest suggests protein channels are closed to conserve energy; however, ATPases work concomitantly in the same membranes with these channels to maintain important ion gradients. One possible mechanism causing downregulation of ion pumps and ion leak through channels is by changing membrane lipid composition. Regulation occurs by limiting conformational mobility of integral proteins, whereby components of biological membranes cause rigidity in microdomains where enzymes or protein channels reside, effectively prohibiting catalytic function. Understanding the mechanisms by which membrane-bound ATPases are downregulated will provide key insight into maintenance of ion gradients during aestivation.
The P-type family of ATPases involves a phospho-intermediate stage of ATP hydrolysis, whereby the terminal phosphoryl group of ATP is transferred to an aspartic acid residue in the active site of the enzyme. Included in this family are type-II P-type ATPases, characterized by their ability to transport low atomic mass cations (H+, Na+, K+, and Mg2+), needed to maintain electrochemical gradients for uptake of nutrients into cells and other functions. These ATPases are well documented as crucial for the survival of all animals and are found in plasma membranes and intracellular membranes. Structurally, all P-type ATPases are made up of the catalytic α-subunit and the non-catalytic glycoprotein β-subunit in a 1:1 ratio, though there are several isoforms of each. Furthermore, they are tethered to membranes by protein-lipid interactions and are asymmetric tetramers consisting of 2-4 cooperative α-β subunits, with only the α subunit interacting with membranes. Most importantly, the catalytic a-subunit has 10 transmembrane domains, which can be highly influenced by the composition of its surrounding membrane. This property dictates a plausible means of regulating the enzymes during metabolic depressions associated with membrane lipid alteration.
There are several membrane-bound ATPases that are important consumers of ATP. The magnesium (Mg2+) ATPase (E.C. 126.96.36.199) is typically plasma membrane bound and is involved in extrusion of Mg ions from the cell and is found in both plasma membrane and sarcoplasmic reticulum. Magnesium ions are used as cofactors for many other enzymes including other ATPases. Calcium (Ca2+) concentrations are high both extracellularly (10-3 M) and within the sarcoplasmic reticulum (10-3 M), but low intracellularly (10-6 M). Ca2+-ATPase (E.C. 188.8.131.52) in both the plasma and sarcoplasmic reticulum membranes maintains low intracellular calcium concentrations and sequesters large amounts in the sarcoplasmic reticulum for use in muscle contraction.
The most widely studied membrane ATPase, Na+/K+-ATPase (E.C. 184.108.40.206), maintains the electrochemical gradients of both sodium and potassium ions across the plasma membrane. This enzyme is present in rat cardiomyocyte sarcoplasmic reticulum, but has not been reported in sarcoplasmic reticulum of any other species or tissue. Another ATPase known for high consumption of cellular ATP, and therefore a potential target for downregulation during metabolic depression is the P-type H+-ATPase (E.C. 220.127.116.11). This enzyme differs from the vacuolar (V-type) proton ATPase in location and structure, and actively facilitates the efflux of H+ ions from the cell when embedded in plasma membrane. Additionally, proton movement via this enzyme drives the influx of Na+ ions into cells and cellular compartments. Measurements of activities of these membrane-bound ATPases and compositions of their surrounding lipid milieu can be used to assess the extent to which their downregulation contributes to metabolic depression in aestivation.