STIMULATION OF CA2-PUMPING ATPASE ACTIVITY IN CARROT PLASMA-MEMBRANE BY CALMODULIN()

Activity of Ca2+-pumping ATPase located at the plasma membrane of cult ured carrot cells was appreciably stimulated when the enzyme was assoc iated with calmodulin (CAM). Affinity of the ATPase toward the substra tes was increased by the addition of CAM, and K(m)s of the enzyme for Ca2+ were estimated to be 11.4 and 0.7 mu M in the absence and presenc e of CAM, respectively, while the values for ATP decreased only slight ly (914 and 670 mu M). In contrast, relative V values of the enzyme we re essentially not changed even after the addition of CAM. This ATPase was capable of utilizing several ATP analogues, such as GTP, ITP, UTP and CTP, instead of ATP, and Ca2+-transport driven by these alternati ve substrates was also stimulated by CAM. The molar ratio of Ca2+-tran sport and ITP hydrolysis of the enzyme reaction was estimated to be 2: 1. These observations suggested that CAM-induced stimulation of Ca2+-A TPase activity at the plasma membrane of cultured carrot cells is main ly due to the increase in the affinity of the enzyme toward Ca2+ by as sociation with the modulator protein, and the hydrolysis of 1 mol of G TP by the enzyme action results in the transport of 2 mol of Ca2+ acro ss the membrane. (C) 1997 Elsevier Science Ltd. All rights reserved.