Characterization of a calmodulin-binding transporter from the plasma membrane of barley aleurone

<jats:p> We have used <jats:italic>Arabidopsis</jats:italic> calmodulin (CaM) covalently coupled to horseradish peroxidase to screen a barley aleurone cDNA expression library for CaM binding proteins. The deduced amino acid sequence of one cDNA obtained by this screen was shown to be a unique protein of 702 amino acids with CaM and cyclic nucleotide binding domains at the carboxyl terminus and high similarity to olfactory and K <jats:sup>+</jats:sup> channels. This cDNA was designated HvCBT1 ( <jats:italic>Hordeum vulgare</jats:italic> CaM binding transporter). Hydropathy plots of HvCBT1 showed the presence of six putative transmembrane domains, but sequence alignment indicated a pore domain that was unlike the consensus domains in K <jats:sup>+</jats:sup> and olfactory channels. Expression of a subclone of amino acids 482–702 in <jats:italic>Escherichia coli</jats:italic> generated a peptide that bound CaM. When a fusion protein of HvCBT1 and green fluorescent protein was expressed in barley aleurone protoplasts, fluorescence accumulated in the plasma membrane. Expression of HvCBT1 in the K <jats:sup>+</jats:sup> transport deficient <jats:italic>Saccharomyces cerevisiae</jats:italic> mutant CY162 showed no rescue of the mutant phenotype. However, growth of CY162 expressing HvCBT1 with its pore mutated to GYGD, the consensus sequence of K <jats:sup>+</jats:sup> channels, was compromised. We interpret these data as indicating that HvCBT1 acts to interfere with ion transport. </jats:p>