Polyols that Accumulate in Renal Tissue Uncouple the Plasma Membrane Calcium Pump and Counteract the Inhibition by Urea and Guanidine Hydrochloride

Abstract

Sorbitol and mannitol, two stereoisomeric osmolytes, inhibit the ATP-dependent Ca2+ transport in inside-out vesicles derived from basolateral membranes from kidney proximal tubules. This inhibition (I0,5 = 400 and 390 mᴍ respectively) cannot be attributed to an in­ crease in Ca2+ permeability, since the rate of EGTA-stimulated Ca2+ efflux from preloaded vesicles is not modified by these osmolytes. In the presence of 1 ᴍ sorbitol or mannitol, Ca2+ uptake is inhibited by 70 and 75%, respectively. Since the Ca2+-stimulated ATPase activity is unaffected, sorbitol and mannitol uncouple the Ca2+ transport from the ATPase activity. The inhibition of Ca2+ transport by these osmolytes is reversible, since the inhibition dis­appears when the vesicles are preincubated with 1 ᴍ sorbitol or mannitol and then diluted 25-fold in reaction medium to measure Ca2+ accumulation. On the other hand, these osmo­lytes protect the (Ca2+ +Mg2+)ATPase from the inhibition of Ca2+ transport and ATPase activity by urea and guanidinium. These data suggest that the high concentrations of polyols that renal cells accumulate during antidiuresis, may regulate Ca2+ transport across the plasma membrane. In addition, polyols may protect the (Ca2+ + Mg2+) ATPase from the deleterious structural effects of urea, a compound that also accumulates during antidiuresis.

Protection against thermal denaturation by trehalose on the plasma membrane H+‐ATPase from yeast

“…It has been demonstrated that some osmolytes protect against the effects of urea and guanidinium chloride on the function of different enzymes [13,14,37–40]. We have previously shown that the inactivation of plasma membrane H + ‐ATPase by guanidinium chloride is not protected by trehalose [14]; however, when the solubilized enzyme was reconstituted in artificial liposome, the functional stability was increased, although the presence of trehalose did not contribute to further increase it [14].…”

“…Carbohydrates and polyols can modulate the catalytic cycle of ion‐transporting ATPases, the so‐called P‐ATPases. These effectors decrease the K m for P i in the phosphorylation reaction catalyzed by the (Ca 2+ + Mg 2+ )ATPase from sarcoplasmic reticulum [52], uncouple Ca 2+ transport from ATP hydrolysis [13,53] and increase the rate of ATP–P i exchange catalyzed by the (Ca 2+ + Mg 2+ )ATPase from renal plasma membrane [37,38]. All these reactions are stimulated by a decrease in water activity and in protein solvation and are favored by the stabilization of E 2 conformation of these enzymes [52,54].…”

“…Organisms and cellular systems are required to adapt to stress conditions such as high temperature, often responding by accumulating organic solutes such as sugars, polyols, amino acids or methylamines [5,10–15]. This accumulation is associated with the effectiveness of these osmolytes in minimizing protein denaturation and membrane damage under stress conditions [16–21].…”

Carbohydrate Protection of Enzyme Structure and Function against Guanidinium Chloride Treatment Depends on the Nature of Carbohydrate and Enzyme

“…Recently, it has been shown that many osmolytes, including carbohydrates and polyols, modulate the functions of different enzymes (Vieyra et al, 1989Chini et al, 1991;, 1995a, and, in addition, can preserve the function and/or structure of enzymes against chemical and physical stresses Meyer-Fernandes, 1994, 1996). It has been shown that among the carbohydrates tested, trehalose is the most effective, protecting yeast cytosolic enzymes against thermal inactivation ), suggesting why yeasts accumulate trehalose but not other carbohydrates when submitted to a heat shock (Wiemken, 1990;De Virgilio et al, 1991Hottiger et al, 1994;Meyer-Fernandes, 1994, 1996).…”

Betaine protects urea‐induced denaturation of myosin subfragment‐1

“…Often, they are further classified as 'compatible' or 'counteracting' on the basis of their ability to affect the functional activity of proteins. Compatible osmolytes increase protein stability against denaturation with little or no effect on their function [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20], whereas counteracting osmolytes have, in addition, the ability to offset the deleterious effects of denaturants on the catalytic activity [1,3,[6][7][8][11][12][13][17][18][19][20][21][22][23][24][25]. The counteracting solute effect can be viewed as a situation in which the tendency of a denaturant to unfold the protein structure is counterbalanced by a tendency of the solute to minimize the protein surface area in contact with water [18][19][20]25].Myosin (EC 3.6.4.1) belongs to the restricted family of motor proteins that transform the chemical energy of ATP into mechanical work.…”