Expression of the cloned subunits of Escherichia coli transhydrogenase from separate replicons.

ABSTRACT

The pntA and pntB genes of Escherichia coli, encoding the alpha- and beta-subunits of the pyridine nucleotide transhydrogenase, were cloned individually in two different compatible plasmids into Escherichia coli mutants lacking transhydrogenase activity. Energy-linked and non-energy-linked transhydrogenase activities were produced only in cells carrying both plasmids thus showing that the products of both genes are required for the formation of an active enzyme. ATP-energized transhydrogenase activity was not increased in cells containing amplified levels of the transhydrogenase when the cell membrane ATPase was also amplified. It is suggested that the excess transhydrogenase is effectively uncoupled from the ATPase by compartmentalization in the cell.