CDP phosphotransferase activity in spinach intact chloroplasts: Possible involvement of nucleoside diphosphate kinase II

Nucleotides formation after addition of [γ32P]-ATP has been analysed in isolated chloroplasts in the presence of exogenous CDP, UDP and GDP. The highest level of phosphotransfer was observed on CDP and UDP after 10 min incubation. Interestingly, the phosphorylation increase of chloroplastic CDP in organello correlated with the time-dependent dephosphorylation of a 18-kDa polypeptide, thereby indicating that CDP, the major endogenous phosphorylated NDP, is likely to be a potent in vivo substrate for this phosphoprotein. The 18-kDa polypeptide was immunoprecipitated with antibodies directed against human nucleoside diphosphate kinase (NDPK) A/B and spinach NDPK-II, both belonging to the ubiquitous family of NDPKs (EC 2.7.4.6). Using recombinant NDPK-II, we could not show a preference for CDP in vitro, suggesting either that CDP is the most available NDPK-II substrate in intact chloroplasts or a chloroplastic factor modulates the enzyme affinity for nucleoside diphosphate substrates in vivo.