Expression of the vhb gene encoding hemoglobin from Vitreoscilla stercoraria in several organisms, clearly enhances oxygen-dependent product formation. In a previous work, we expressed the vhb gene that encodes hemoglobin from V. stercoraria in Amycolatopsis mediterranei, resulting in an increase (oxygen-dependent formation) in rifamycin B production. In the present work, we first confirm; by heterologous expression in Escherichia coli, that rif-orf5 from the rifamycin biosynthetic gene cluster, really encodes a cytochrome P450 enzyme, which is the key step for oxygen incorporation in the final biosynthetic product. Likewise, we fused rif-orf5 to the vhb gene, as part of a genetic engineering strategy. The fused genes were used to generate an Amycolatopsis mediterranei transformant (Msb-HbCYP5). Interestingly, the fermentation of Msb-HbCYP5 manifested 1.5-fold higher rifamicin B production than the transformant with only the hemoglobin gene, and 2.2-fold higher than the parental strain.
Keywords: Actinomycetes; Amycolatopsis; Cytochrome P450; Hemoglobin; Rifamycin; Vitreoscilla.