Downloaded by COLUMBIA UNIV on July 27, 2012 | http://pubs.acs.org Publication Date: May 5, 1995 | doi: 10.1021/ba-1995-0250.ch019
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Electric and Magnetic Field Signal Transduction in the Membrane Na /K -adenosinetriphosphatase +
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Martin Blank Department of Physiology and Cellular Biophysics, Columbia University, New York, NY 10032 +
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The Na /K -adenosinetriphosphatase (Na /K -ATPase), the ion-pump enzyme located in cell membranes, is a well-defined protein whose properties can be used to study the mechanism of electromag netic (EM)fieldinteraction with biological systems. Enzyme activity is normally inhibited by electricfieldsand stimulated by magnetic fields, but bothfieldscause large increases in enzyme activity when the initial (basal) activity of the enzyme is greatly reduced by aging, by lowering temperature, or by inhibitors. The opposing effects un der optimal conditions suggest different charge movements in differ ent parts of the enzyme. Electricfieldsact as if they increase ion binding at the enzyme surface, whereas magneticfieldsappear to affect charges within the protein (e.g., charge transfer due to the ATPase reaction). The similar effects under suboptimal conditions suggest that the two different charge movements are coupled. The coordinated changes in charge density caused by bothfieldssuggest ways in which EMfieldsaffect membrane proteins ingeneral.They also suggest ways in which the Na/K-ATPase may function as an ion pump. +
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NTEREST IN BIOLOGICAL EFFECTS of electromagnetic (EM) fields has been driven by epidemiological studies that report increased cancer risk associated with exposure. Changes in biosynthesis in cells following EM interaction sug0065-2393/95/0250~O339$12.00/0 ©1995 American Chemical Society
In Electromagnetic Fields; Blank, M.; Advances in Chemistry; American Chemical Society: Washington, DC, 1995.
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ELECTROMAGNETIC FIELDS
gest biochemical pathways that could be responsible for the association, but that area of research has not clarified the initial signal transduction step. We do not know how weak E M fields interact with molecules or how molecular reactions initiate complex cellular processes. To approach the problem of transduction of E M field exposures into molecular changes, that is, which charges are affected and how and when they move in the molecule, we have studied the effects of electric and magnetic fields on the enzyme activity of a well-defined membrane protein, Na /K -adenosinetriphosphatase (Na /K -ATPase). +
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Na/Jt-ATPase: The Ion-Pump Enzyme Downloaded by COLUMBIA UNIV on July 27, 2012 | http://pubs.acs.org Publication Date: May 5, 1995 | doi: 10.1021/ba-1995-0250.ch019
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The Na /K -ATPase is a ubiquitous transport enzyme whose structure and en zymatic properties (e.g., ion binding sites, ion activation, inhibitors, and tem perature dependence) are well known. Its properties were first described in 1957 by Skou (1), who demonstrated activation by cations. The molecular structure and many properties of the enzyme have been reviewed in monographs by Lauger (2) on electrogenic ion pumps and by Tonomura (3) on energy-trans ducing ATPases. The Na /K -ATPase is composed of two polypeptide chains (a and β) that extend through the membrane, and the catalytic activity (the 110-kDa α-chain) is directly affected by the ionic concentrations in contact with the two sides of the enzyme. The enzyme is activated when N a ions bind to the inside surface and K ions to the outside surface. Research on red blood cells established the stoichiometry of the pump (three N a ions out and two K ions in for each A T P split) and its reversibility (i.e., ions flowing down gradients exceeding normal reverse the action of the enzyme and cause the synthesis of A T P from precur sors). When the purified molecule is introduced into an artificial bilayer (4), it forms a gated ion channel. The enzyme is phosphorylated in the presence of N a ions and dephosphorylated in the presence of K ions. The two conformational states are E i when N a ions (and ATP) bind from the inside, and E when K ions bind from the outside. The ion binding sites are not fully accessible to exchange with other ions in the medium in the two conformational states, and this property is referred to as occlusion. Also, some transitions between conformational states are volt age-sensitive (5). These studies have led to the formulation of cyclic schemes that alternate the two conformational states to link ion transport to N a / K ATPase function. Post (6) has described the enzyme as an oscillator and has suggested that it can be forced between conformational states by imposing oscil lating mechanical or electrical stimuli. Post's ideas about the enzyme mechanism were stimulated by the work of Tsong and co-workers (7-11), who described a ouabain-sensitive accumulation of rubidium and secretion of sodium from red blood cells stimulated by alternating currents. The Tsong group suggests that the electric field interacts directly with the enzyme, driving it through conforma tional states that cause ion movements. This interaction has been referred to as the electroconformational coupling model. +
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In Electromagnetic Fields; Blank, M.; Advances in Chemistry; American Chemical Society: Washington, DC, 1995.
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BLANK
EMF Signal Transduction in Membrane Na+/K -ATPase
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Effects ofAC Electric Fields on Na/it-ATPase Activity We have characterized (12-17) the effects of alternating current (AC) electric fields on Na /K -ATPase activity. A C normally decreases the rate of A T P split ting in correlation with the level of ion activation, but it can increase activity when the basal enzyme activity is greatly reduced by low temperature, by inhibi tors (ouabain), or by changing the concentrations of activating cations. Figure 1 shows how the effect of an electric field varies with enzyme activity. Above an enzyme activity of 0.05-0.1 μπιοί phosphate-mg protein -min , the ratio of enzyme activity with A C to enzyme activity without A C (E/A) is 1. The low values of enzyme activity were obtained by using different concentrations of ouabain, and similar effects were obtained by using lowered temperatures. Because enzyme activity varies with ion concentration, by increasing at low activity and decreasing (or reaching a plateau) at high ion concentration, both inhibition and stimulation by A C electric fields suggest a field-dependent increased binding of activating cations (12). A n increase in binding has opposite effects in different ranges of enzyme activity (1). A t optimal activity, increased ion concentration lowers activity. When basal activity is lowered by inhibitors or low temperature, increased ion concentration increases enzyme activity. In studies in which enzyme activity varies with Na/K ratio, the effect of A C corre lates directly with basal enzyme activity. Independent evidence for an inhibitory effect of electric fields on N a / K ATPase activity was published by Green et al. (18), who reported an almost twofold increase in enzyme content, measured by ouabain binding, as a result of stimulation at 10 H z for several days. Because enzyme is synthesized in re+
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Downloaded by COLUMBIA UNIV on July 27, 2012 | http://pubs.acs.org Publication Date: May 5, 1995 | doi: 10.1021/ba-1995-0250.ch019
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