Most Streptomyces strains are equipped with only the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway for the formation of isopentenyl diphosphate. In addition to this pathway, some Strepromyces strains have the mevalonate pathway to produce terpenoid antibiotics. We have previously shown that a gene cluster for biosynthesis of terpentecin, a diterpene antibiotic, was located in adjacent the mevalonate pathway gene cluster. In this study, a mevalonate pathway gene cluster was cloned from Actinoplanes sp. strain A40644, an isoprenoid antibiotic BE-40644 producer, to examine whether the mevalonate pathway genes and isoprenoid biosynthetic genes are clustered in genomic DNA. By sequencing flanking regions a probable BE-40644 biosynthetic gene cluster was found in the downstream region of the mevalonate pathway gene cluster. Heterologous expression of a 9-kb fragment confirmed that a set of the BE-40644 biosynthetic genes was involved in the fragment. This result suggested that the presence of the mevalonate pathway might be a good landmark to detect the production of isoprenoid compounds by actinomycetes.