The ribosome is essential for protein synthesis and targeting its function serves as a platform for many therapies. Resistance gene-guided genome mining for a priori target identification has emerged as a useful workflow in natural product drug discovery, allowing for identification of a compound's target before the time-consuming work of isolation and characterization is performed. In this work, we identify the previously unknown lig biosynthetic gene cluster (BGC) which contains a predicted resistance gene that shares sequence identity with the gene for human ribosomal protein RPL10. Through both heterologous expression and in situ BGC engineering, we identify ligustrone A (1) as this BGC's product. Utilizing chemical genetics and chemoproteomics, we validate that 1 engages RPL10. Taken together, via resistance gene-guided mining we both discover a new target for a known natural product and elucidate its biosynthesis.