The Calcium-Binding Activity of a Vacuole-Associated, Dehydrin-like Protein Is Regulated by Phosphorylation

• Published in: Plant physiology (Bethesda) Vol. 130; no. 2; pp. 675 - 687
• Main Authors: Bruce J. Heyen, Muath K. Alsheikh, Smith, Elizabeth A., Carl F. Torvik, Seals, Darren F., Randall, Stephen K.
• Format: Journal Article
• Language: English
• Published: Rockville, MD American Society of Plant Biologists 01.10.2002; American Society of Plant Physiologists
• Subjects: Acid Phosphatase - metabolism; Alkaline Phosphatase - metabolism; Amino Acid Sequence; Amino acids; Antibodies; Apium graveolens; Apium graveolens - genetics; Apium graveolens - metabolism; Biological and medical sciences; Blotting, Western; Calcium; Calcium - pharmacology; Calcium binding proteins; Calcium-Binding Proteins - drug effects; Calcium-Binding Proteins - genetics; Calcium-Binding Proteins - metabolism; Celery; Cell biochemistry; Cell Biology and Signal Transduction; Cell physiology; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Gels; Heat treatment; Molecular Sequence Data; Phosphatases; Phosphorylation; Phosphorylation - drug effects; Phosphotransferases - metabolism; Plant physiology and development; Plant Proteins - genetics; Plant Proteins - metabolism; Sequence Homology, Amino Acid; Vacuoles; Vacuoles - metabolism; Tonoplast; Phosphorylation; Purification; Calcium; Apium graveolens; Vacuole; Biological activity; Characterization; Binding protein; Regulation(control); Vegetable crop; Dicotyledones; Angiospermae; Spermatophyta; Umbelliferae; Localization
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.002550

A vacuole membrane-associated calcium-binding protein with an apparent mass of 45 kD was purified from celery (Apium graveolens). This protein, VCaB45, is enriched in highly vacuolate tissues and is located within the lumen of vacuoles. Antigenically related proteins are present in many dicotyledonous plants. VCaB45 contains significant amino acid identity with the dehydrin family signature motif, is antigenically related to dehydrins, and has a variety of biochemical properties similar to dehydrins. VCaB45 migrates anomalously in sodium dodecyl sulfate-polyacrylamide gel electrophoresis having an apparent molecular mass of 45 kD. The true mass as determined by matrix-assisted laser-desorption ionization time of flight was 16.45 kD. VCaB45 has two characteristic dissociation constants for calcium of 0.22 ± 0.142 mM and 0.64 ± 0.08 mM, and has an estimated 24.7 ± 11.7 calcium-binding sites per protein. The calcium-binding properties of VCaB45 are modulated by phosphorylation; the phosphorylated protein binds up to 100-fold more calcium than the dephosphorylated protein. VCaB45 is an "in vitro" substrate of casein kinase II (a ubiquitous eukaryotic kinase), the phosphorylation resulting in a partial activation of calcium-binding activity. The vacuole localization, calcium binding, and phosphorylation of VCaB45 suggest potential functions.