An AtCRK1 [Arabidopsis thaliana CDPK (Ca2+-dependent protein kinase)-related protein kinase 1] has been characterized molecularly and biochemically. AtCRK1 contains the kinase catalytic domain and a CaM (calmodulin)-binding site. Our results demonstrated thatAtCRK1couldbindCaMinaCa2+-dependent manner. This kinase phosphorylated itself and substrates such as histone IIIS and syntide-2 in a Ca2+-independent manner and the activitywas stimulated by several CaM isoforms through its CaMbinding domain. This domain was localized within a stretch of 39 amino acid residues at positions from 403 to 441 with Kd=67 nM for CaM binding. However, the stimulation amplification of the kinase activity of AtCRK1 by different CaM isoforms was similar.