Molecular and biochemical characterization of a calcium/calmodulin-binding protein kinase from rice

A Ca²⁺/calmodulin (CaM)-binding protein kinase from rice (Oryza sativa), OsCBK, has been characterized that lacks Ca²⁺-binding EF hands and has Ca²⁺/CaM-independent autophosphorylation and substrate-phosphorylation activity. OsCBK has all 11 subdomains of a kinase catalytic domain and a putative CaM-binding domain, and shares high identity with Ca²⁺-dependent-protein-kinase ('CDPK')-related protein kinases in plants. OsCBK bound CaM in a Ca²⁺-dependent manner as previously reported for Ca²⁺/calmodulin-dependent protein kinases in animals, but autophosphorylation and phosphorylation of histone IIIs were Ca²⁺/CaM-independent. Surface plasmon resonance analysis showed that OsCBK specifically bound CaM with high affinity (K_D = 30nM). Capillary electrophoresis showed that phosphorylation of OsCBK occurred on serine and threonine residues. These data show that OsCBK is a serine/threonine protein kinase that binds Ca²⁺/CaM, but whose enzymic activity is independent of Ca²⁺/CaM. In situ hybridization showed that OsCBK is expressed in reproductive and vegetative tissues of rice and shows temporal and spatial changes during plant growth and development. OsCBK is highly expressed in zones of cell division and it is particularly abundant in sporogenous cells of the anther at meiosis.