Sarcoplasmic membrane calcium transport

Pathogenesis of MH is not completely understood. Skeletal muscle, however, is the one tissue in MH with proven abnormalities, and it is further thought that the basic defect that causes the syndrome lies in the calcium regulation system found within the myoplasm. For example, calcium transport function appears to be decreased in the sarcoplasmic reticulum, mitochondria, and sarcolemma. Thus, the suggestion has been made that MH is characterized by a generalized membrane defeet. 

ATP is used not only to power muscle contraction, but also to re-establish the resting state of the cell. At the end of the contraction cycle, calcium must be transported back into the sarcoplasmic reticulum, a process which is ATP driven by an active pump mechanism. Additionally, an active sodium-potassium ATPase pump is required to reset the membrane potential by extruding sodium from the sarcoplasm after each wave of depolarization. When cytoplasmic Ca2- falls, tropomyosin takes up its original position on the actin and prevents myosin binding and the muscle relaxes. Once back in the sarcoplasmic reticulum, calcium binds with a protein called calsequestrin, where it remains until the muscle is again stimulated by a neural impulse leading to calcium release into the cytosol and the cycle repeats. 

The identification of the sarcoplasmic reticulum membranes as the intracellular calcium transport system involved in the regulation of muscle activity took considerable time and occurred at first quite fortuitously. Kielley and Meyerhof40, not aware of the existence of the sarcoplasmic reticulum, characterized it biochemically as an... 

The low lipid-protein ratio of 0.5 together with the size of the sarcoplasmic vesicles implies that only approximately 30% of their membranes can be occupied by a regular lipid bilayer structure. Consequently, a large fraction of the membrane protein must interrupt the lipid bilayer and reach throughlt. The fact that only one polypeptide chain constitutes the structural unit of the calcium transport protein strong-... 

It was very early recognized that the calcium transport and the calcium-dependent ATPase could simultaneously be blocked by thiol reagents26). In contrast to various other thiol containing enzymes the activities of the sarcoplasmic reticulum membranes cannot be restored when the blocking agents are removed. 

In contrast to the hydrolysis and synthesis of ATP connected with proton translocation in mitochondria, chloroplasts and bacterial membranes, the energy linked movement of calcium ions gives rise to the appearance of an acid-stable phosphorylated intermediate in the membranes. A cation specific phosphorylation also occurs in the membranes of the sodium potassium transport system183. However, due to the inability to correlate phosphorylation and ion movement in the latter membranes, membrane phosphorylation has been questioned as being a step in the reaction sequence of ion translocation184,18s. Solely the sarcoplasmic calcium transport system allows to correlate directly and quantitatively ion translocation with the phosphoryl transfer reactions. 

The transporting protein in the sarcoplasmic membrane can be phosphorylated by ATP as well as by inorganic phosphate (cf.2,174 ). In the forward running mode of the pump, i. e. when the calcium pump accumulates calcium and concomitantly hydrolyzes ATP, the terminal phosphate residue of ATP is transferred to the transport protein. The reaction depends on the presence of calcium ions in the external medium. In the reverse mode of the pump inorganic phosphate is incorporated into the transport protein. This reaction is inhibited when calcium ions are present in the external medium,... 

Martonosi, A., Boland, R., and Halpin, R. A. The biosynthesis of sarcoplasmic reticulum membranes and the mechanism of calcium transport. Cold Spring Harbor Symp. Quant. Biol. 37, 455-468 (1972). 

Schematic diagram of a cardiac muscle sarcomere, with sites of action of several drugs that alter contractility (numbered structures). Site 1 is Na+/K+ ATPase, the sodium pump. Site 2 is the sodium/calcium exchanger. Site 3 is the voltage-gated calcium channel. Site 4 is a calcium transporter that pumps calcium into the sarcoplasmic reticulum (SR). Site 5 is a calcium channel in the membrane of the SR that is triggered to release stored calcium by activator calcium. Site 6 is the actin-troponin-tropomyosin complex at which activator calcium brings about the contractile interaction of actin and myosin.

Relaxation of the muscle is brought about by removal of the ionic calcium from the sarcoplasm. This calcium is transported across the membrane of the sarcoplasmic reticulum, in an energy requiring process. In addition to the calcium pumping ATPase, the sarcoplasmic reticulum also contains a calcium binding protein called calsequestrin (Section 4.3.3). Some of the calcium segregated by the sarcoplasmic reticulum is apparently bound to this protein within the lumen of the sarcoplasmic reticulum. As sequestration of calcium ions into sarcoplasmic reticulum proceeds, more calcium ions dissociate from their binding sites on troponin C, re-... 

Biochemical alterations have been found in fragmented sarcoplasmic reticulum isolated from dystrophic human, mouse and chicken muscle. Alterations in calcium transport, ATP hydrolysis and phosphoenzyme formation have been reported. Some of these biochemical alterations in the dystrophic sarcoplasmic reticulum are suggested to be due to alterations of the lipid environment of these membranes it has been suggested that the cholesterol content of dystrophic sarcoplasmic reticulum is elevated [182-187]. 

The second domain, where significant progress has been achieved is that of time-resolved studies on membrane processes during active transport. So far this type of studies has been restricted to the calcium transport system of sarcoplasmic reticulum membranes, but the work shows the great potential of such studies also for other functional membrane species, and shall, therefore, be reviewed here in some detail. 

The present review deals with calcium transport and the accompanying enzymatic reactions displayed by the sarcoplasmic reticulum membranes. Subjects such as the molecular organization of the membranes, transport energetics, activity modulation, the development of the calcium transport system and its role in the regulation of muscular activity could not be considered. These problems have been partially reviewed previously (cf. [1-6]). 

In the presence of phosphate donating or accepting reactants, the translocation of calcium ions across the sarcoplasmic membranes is linked with phosphoryl transfer reactions leading to the phosphorylation of the transport protein. During calcium accumulation, the terminal phosphate group of ATP or of the other phosphate donors is rapidly transferred to the transport protein from which it is subsequently liberated by hydrolytic cleavage. The phosphoryl group in the protein is acid-stable and can therefore be stabilized in acidic quench media [112-114]. 

When the sarcoplasmic calcium transport system operates in the reverse mode and synthesizes ATP from ADP and inorganic phosphate during calcium release, inorganic phosphate reacts with the transport protein also leading to the formation of a phosphoprotein [115 -117]. This reaction also requires ionized magnesium but is suppressed when the concentration of ionized calcium in the medium exceeds 10 /xM. In the transport protein of the sodium-potassium system, analogous cation dependent phosphoryl transfer reactions take place. It is difficult, however, to directly correlate phosphorylation and ion movement in these membranes. 

Cholesterol, in various proportions, is a natural constituent of mammalian plasma membranes but, if the proportion is allowed to increase, membrane function is usually diminished. Thus the membrane that surrounds the sarcoplasmic reticulum vesicles in muscle progressively loses the calcium-transporting function of its ATPase when cholesterol starts replacing the phospholipids. Similarly, ox-heart mitochondria, when exposed to cholesterol, progressively lose the activity of ATPase, succinate dehydrogenase, and /3-hydroxybutyrate dehydrogenase (Warren et aL, 1975). 

During development of chicken skeletal muscle cells, the marked increase in Ca transport of sarcoplasmic reticulum, observed both in vivo and in vitro systems, can be interpreted mainly as the result of an increase in the concentration of Ca transport ATPase. Changes in the fatty acid composition of muscle membranes developed in vivo occur with a balance between chain length and unsaturation, without affecting significantly their Ca permeability. In cultured muscle cells their fatty acid composition can be manipulated to a great extent by lipid supplementation of the culture medium. The effects of these in vitro modifications in lipid composition on the calcium transport function of muscle membranes should be investigated. 

MacLennan, D. H., and Holland, P. C., 1976, The calcium transport ATPase of sarcoplasmic reticulum, in The Enzymes of Biological Membranes, Vol. 3, Membrane Transport (A. Martonosi, ed.), pp. 221-259, Plenum Press, New York. 

Mehorta and coworkers (1989) observed that isolated fractions of brain and heart cells from rats orally administered 0.5-10 mg endrin/kg showed significant inhibition of Ca+2 pump activity and decreased levels of calmodulin, indicating disruption of membrane Ca+2 transport mechanisms exogenous addition of calmodulin restored Ca+2-ATPase activity. In vitro exposure of rat brain synaptosomes and heart sarcoplasmic reticuli decreased total and calmodulin-stimulated calcium ATPase activity with greater inhibition in brain preparations (Mehorta et al. 1989). However, endrin showed no inhibitory effects on the calmodulin-sensitive calcium ATPase activity when incubated with human erythrocyte membranes (Janik and Wolf 1992). In vitro exposure of rat brain synaptosomes to endrin had no effect on the activities of adenylate cyclase or 3, 5 -cyclic phosphodiesterase, two enzymes associated with synaptic cyclic AMP metabolism (Kodavanti et al. 1988). 

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