Identification of a calmodulin‐stimulated (Ca2++ Mg2+)‐ATPase in a plasma membrane fraction isolated from maize (Zea mays) leaves

Abstract

Plasma membranes were isolated from light‐grown, 14‐day‐old maize leaves (Zea mays L. cv. Golden Cross Bantam) using aqueous two‐phase partitioning. The plasma membrane (PM) fraction contained < 0.3% of the total chlorophyll, < 0.2% of the mitochondrial marker enzyme activity, minimal contamination by endomembranes and 34% of the total PM. A calmodulin‐stimulated (Ca2++ Mg2+)‐ATPase was identified in the PM‐enriched fraction. The Ca2++ calmodulin stimulation was dependent on Mg2+, saturated at ca 25 μM total Ca2+, had a pH maximum at 7.2 and was maximally stimulated by 600 nM bovine brain calmodulin. The stimulation was not greatly affected by the anion present and showed a divalent cation specificity of Ca2+ > Sr+2 ± Mn+2 > Co2+± Cu2+ > Ba2+. The napthalenesulfonamide W7, an antagonist of calmodulin action, completely inhibited the calmodulin stimulation at 175 μM, while the less active analogue W5 was ineffective at this concentration. La3+, an inhibitor of PM Ca2+ transport, showed a 50% inhibition of calmodulin‐stimulated ATPase activity at ca 200 μM. Taken as a whole, these data demonstrate the presence of a calmodulinstimulated, (Ca2++ Mg2+)‐ATPase on the cytoplasmic surface of the plasma membrane of maize leaf cells.

A Single Gene May Encode Differentially Localized Ca 2+ -ATPases in Tomato

“…These data are consistent with the lack of a calmodulin binding domain in the deduced LCA amino acid sequence (Wimmers et al, 1992). In previous studies, calmodulin sensitivity was reported for the PM Ca2+-ATPases from maize (Robinson et al, 1988;Briars and Evans, 1989), red beet roots (Williams et al, 1990), carrot cell cultures (Hsieh et al, 1991), and radish seedlings (Rasi-Caldogno et al, 1992. In contrast, calmodulin had no effect on the PM Ca2+-ATPases from pea roots (Butcher and Evans, 1987) and Commelina communis leaves (Siebers et al, 1990).…”

Subcellular and Tissue Distribution, Partial Purification, and Sequencing of Calmodulin‐Stimulated Ca2+‐Transporting ATPases from Barley (Hordeum Vulgare L.) and Tobacco (Nicotiana Tabacum)

“…A 140-kDa protein that cross-reacts with antibodies against the erythrocyte Caz+ pump was reported to be present in maize coleoptiles (Briars et al, 1988), and the activity of a PM-located Ca2+ ATPase has been studied in maize leaves (Robinson et al, 1988), radish seeds (Rasi Caldogno et al, 1989, 1995, and Commelina communis leaves (Graf and Weiler, 1990). An intracellular CaM-stimulated Ca2+ transporter, presumably in the ER, was reported for maize roots (Brauer et al, 1990), carrot suspension cultures (Hsieh et al, 1991), cauliflower inflorescences (Askerlund and Evans, 1992) and barley aleurones (Gilroy and Jones, 1993).…”

“…The membranes were collected after centrifuging for IS min in an Eppendorf centrifuge. The pellet was washed with cold distilled water and subjected to acidic SDS/PAGE according to the method of Sarkadi et al (1986). The dried gel was exposed for autoradiography.…”

A Plasma Membrane Protein from Zea mays Binds with the Herbivore Elicitor Volicitin

“…Membrane fractions enriched in plasma membrane material were isolated from Z. mays leaves following the protocol of Robinson et al (1988), with all subsequent operations conducted at 48C. Described briefly, leaves were homogenized in 15 mM Tris-HCl, pH 7.8, with 1 mM EDTA, 3 mM DTT, 0.5 M sucrose, and 0.6% polyvinylpyrrolidone in a 1:4 ratio (g/mL) and filtered through four layers of cheesecloth.…”