A Novel Calmodulin-regulated Ca2+-ATPase (ACA2) from Arabidopsis with an N-terminal Autoinhibitory Domain

• Published in: The Journal of biological chemistry 1998-01, Vol.273 (2), p.1099-1106
• Main Authors: Harper, Jeffrey F., Hong, Bimei, Hwang, Ildoo, Guo, Hong Qing, Stoddard, Robyn, Huang, Jing Feng, Palmgren, Michael G., Sze, Heven
• Format: Article
• Language: English
• Subjects: ACTIVIDAD ENZIMATICA; ACTIVITE ENZYMATIQUE; ADN; Amino Acid Sequence; AMINO ACID SEQUENCES; Arabidopsis - enzymology; ARABIDOPSIS THALIANA; Base Sequence; CA2+-TRANSPORTING ATPASE; CALCIO; CALCIUM; Calcium-Transporting ATPases - antagonists & inhibitors; Calcium-Transporting ATPases - genetics; Calcium-Transporting ATPases - metabolism; CALMODULIN; Calmodulin - metabolism; CALMODULINA; CALMODULINE; CATION; CATIONES; CATIONS; CELL MEMBRANES; CHEMICAL COMPOSITION; CHEMISTRY; CHIMIE; COMPLEMENTARY DNA; COMPLEMENTATION; COMPOSICION QUIMICA; COMPOSITION CHIMIQUE; DNA; DNA, Complementary; ENZYMIC ACTIVITY; ESTIMULO; FLEUR; FLORES; FLOWERS; GENBANK/AF025842; Genetic Complementation Test; GENETICA; GENETICS; GENETIQUE; HIDROLASAS; HYDROLASE; HYDROLASES; IMMUNOCYTOCHEMISTRY; IMMUNOHISTOCHEMISTRY; IMMUNOLOGIE; IMMUNOLOGY; INMUNOLOGIA; ION; ION TRANSPORT; IONES; IONS; Life Sciences (General); MEMBRANAS CELULARES; MEMBRANE CELLULAIRE; MOLECULAR SEQUENCE DATA; MUTACION; MUTATION; NUCLEOTIDE SEQUENCE; Plant Proteins - antagonists & inhibitors; Plant Proteins - genetics; Plant Proteins - metabolism; PLASMA MEMBRANES; Protein Binding; PYROPHOSPHATASES; QUIMICA; RACINE; RAICES; ROOTS; SACCHAROMYCES CEREVISIAE; Saccharomyces cerevisiae - genetics; SECUENCIA NUCLEOTIDICA; Sequence Homology, Amino Acid; SEQUENCE NUCLEOTIDIQUE; Space life sciences; STIMULI; STIMULUS
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.2.1099

To study transporters involved in regulating intracellular Ca2+, we isolated a full-length cDNA encoding a Ca2+-ATPase from a model plant,Arabidopsis, and named it ACA2( ArabidopsisCa2+-ATPase, isoform2). ACA2p is most similar to a “plasma membrane-type” Ca2+-ATPase, but is smaller (110 kDa), contains a unique N-terminal domain, and is missing a long C-terminal calmodulin-binding regulatory domain. In addition, ACA2p is localized to an endomembrane system and not the plasma membrane, as shown by aqueous-two phase fractionation of microsomal membranes. ACA2p was expressed in yeast as both a full-length protein (ACA2-1p) and an N-terminal truncation mutant (ACA2-2p; Δ residues 2–80). Only the truncation mutant restored the growth on Ca2+-depleted medium of a yeast mutant defective in both endogenous Ca2+ pumps, PMR1 and PMC1. Although basal Ca2+-ATPase activity of the full-length protein was low, it was stimulated 5-fold by calmodulin (50% activation around 30 nm). In contrast, the truncated pump was fully active and insensitive to calmodulin. A calmodulin-binding sequence was identified within the first 36 residues of the N-terminal domain, as shown by calmodulin gel overlays on fusion proteins. Thus, ACA2 encodes a novel calmodulin-regulated Ca2+-ATPase distinguished by a unique N-terminal regulatory domain and a non-plasma membrane localization.