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The melleolides are a family of structurally and functionally diverse sesquiterpenoids with potential applications as fungicides, antimicrobials, and cancer therapeutics. The initial and terminal steps of the biosynthesis pathway in Armillaria spp. have been characterized, but the intermediate steps are unclear. Biosynthetic gene clusters in A. mellea and A. gallica were shown to encode a terpene cyclase, a polyketide synthase, and four CYP450 monooxygenases. We have characterized CYPArm3, which is responsible for the hydroxylation of Δ‐6‐protoilludene, but the functions of the other CYP450s remain to be determined. Here we describe CYPArm2, which accepts Δ‐6‐protoilludene and 8α‐hydroxy‐6‐protoilludene as substrates. To investigate the products in more detail, we generated recombinant Saccharomyces cerevisiae strains overexpressing CYPArm2 in combination with the previously characterized protoilludene synthase and 8α‐hydroxylase. Using this total biosynthesis approach, sufficient quantities of product were obtained for NMR spectroscopy. This allowed the identification of 8α,13‐dihydroxy‐protoilludene, confirming that CYPArm2 is a protoilludene 13‐hydroxylase.