The inorganic polyphosphate/ATP-dependent NAD kinase (Ppnk) of Mycobacterium tuberculosis was applied to the mass-production of NADP from NAD land inorganic polyphosphate (metaphosphate). When Ppnk purified from recombinant Escherichia coli cells overexpressing the M. tuberculosis Ppnk was used, 30 mM (27 g/l) NADP was produced from 50 mM NAD and 100 mg/ml metaphosphate at 37degreesC and pH 7.0. The recombinant E. coli cells were immobilized in polyacrylamide gel, and treated with acetone to render the cells permeable to substrates and products. When acetone-treated immobilized cells were used, 16 mM (14 g/l) NADP was produced from 50 mM NAD and 100 mg/ml metaphosphate at 37degreesC and pH 7.0. The isolation of NADP formed in the reaction mixture was easy because of the absence of by-products (ATP degradation compounds), and this NADP production system using purified Ppnk or immobilized recombinant E. coli cells expressing Ppnk is thought to be feasible in the production of NADP on an industrial scale.