Vitamin B6 is an essential metabolite that has recently been implicated in defense against cellular oxidative stress. In fungi, the de novo biosynthetic pathway of vitamin B6 involves two genes, PDX1 and PDX2. Here, we report a component of the PDX1/PDX2 vitamin B6 biosynthetic pathway in an arbuscular mycorrhizal (AM) fungus. Using rapid amplification of cDNA ends, we isolated the full-length cDNA of a PDX-like gene, GintPDX1, from Glomus intraradices. GintPDX1 expression was analysed by real-time reverse transcription-polymerase chain reaction (RT-PCR). GintPDX1 activity and function were investigated by heterologous complementation of the yeast strainDeltasnz1, which is deficient in vitamin B6 biosynthesis. Sequence data revealed that GintPDX1 is highly homologous to other identified PDX1 proteins. GintPDX1 restores prototrophy to the vitamin B6 auxotrophic yeast mutant and reverts its superoxide sensitivity. GintPDX1 is expressed throughout the fungal life cycle, with the highest transcription levels found in the intraradical fungal structures. GintPDX1 expression was induced in response to hydrogen peroxide, paraquat and copper. The results demonstrate that AM fungi possess at least one component of the machinery necessary for vitamin B6 biosynthesis. Transcriptional regulation of GintPDX1 suggests a role for vitamin B6 as an antioxidant and modulator of reactive oxygen species in G. intraradices.