Sequence analysis of the lkmB region of the daunorubicin biosynthetic gene cluster of Streptomyces griseus JA3933 revealed two contiguous open reading frames (ORF) in the same orientation, and three ORFs in the opposite orientation together extending over a 4.6 kb region adjacent to a homologue of the S. peucetius dnrJ gene. ORF1 complemented in trans the lkmB mutation, which seems to affect an early step in daunorubicin biosynthesis. Its deduced product showed no similarity to any known enzyme in the databases. The mutation in ORF1 was localised to a C-T transition at position 1172, leading to the change from a glycine to aspartic acid in the deduced protein. The lack of any homology to known polyketide synthesis enzymes indicates a regulatory role for the product of ORF1, despite the ability of lkmB mutants to further metabolise alkanoic acid. The genes of the oppositely oriented cluster seem to be involved in sugar metabolism. The putative ORF3 protein revealed strong homology to eukaryotic acyl CoA dehydrogenases and might encode an enzyme for the oxidoreduction preceding the introduction of the amino group into daunosamine, and the ORF4 protein is homologous to several epimerases, central enzymes in the formation of the L-2,3,6-trideoxy-3-aminohexoses from TDP-D-glucose. ORF5 seems also to be related to enzymes metabolising nucleotide-activated hexoses.