Abstract: This invention relates to a process for the enhancement of naphthalene dioxygenase activity in organisms that have been transformed with DNA encoding the expression of a multiple component naphthalene dioxygenase enzyme. Cells containing the naphthalene dioxygenase enzyme are capable of producing indigo when cultured in the presence of indole. The invention includes procedures to enhance naphthalene dioxygenase activity by transforming a host cell with a DNA sequence that encodes at least two ferredoxin polypeptides for each reductase polypeptide and iron-sulfur polypeptide, by growing transformed organisms in the presence of iron, or by transforming a host cell with a DNA sequence that encodes a ferredoxin analog.

Abstract: Microbial synthesis of indigo dyestuff in indole-free media is disclosed. Indigo production is preferably accomplished by genetic transformation of selected host cells having the capacity to produce and accumulate indole (either as a result of endogenous genomic capacity or genetic transformation) to incorporate the capacity for synthesis of an aromatic dioxygenase enzyme. Growth of transformed cells under suitable conditions facilitates aromatic dioxygenase enzyme catalyzed oxidative transformation of cellular indole, with consequent formation of indigo from the oxidized reaction products. In a highly preferred embodiment, E. coli cells having endogenous indole production capacity are transformed with a DNA expression vector comprising the structural gene for naphthalene dioxygenase, resulting in the microbial synthesis of isolatable quantities of indigo.