Characterization and hetereologous expression of cDNAs encoding two novel closely related Ca2+-binding proteins in Dictyostelium discoideum

During a yeast two hybrid screen of a Dictyostelium cDNA library using the Ca2+-binding protein CBP1 as bait, we isolated a full-length cDNA encoding a novel Ca2+-binding protein (termed CBP4a). The protein is composed of 162 amino acids and contains four consensus EF-hands. PCR amplification of Dic tyastelium genomic DNA using primers specific for the cDNA sequence resulte d in the isolation of a gene encoding a different Ca2+-binding protein of 1 62 amino acids (designated CBP4b) with 90% amino acid sequence identity to CBP4a. Southern blot analysis confirmed the presence of two closely related genes in the Dictyostelium genome. CBP4a and CBP4b mRNAs are expressed at the same stages of development as CBP1 mRNA. In addition, both novel protei ns bind Ca-45(2+) and interact with CBP1 in vitro in a Ca2+ dependent manne r. (C) 2000 Elsevier Science B.V. All rights reserved.