A gene encoding dehydroquinate dehydratase (DHQase) was cloned fromStreptomyces hygroscopicusvar.ascomyceticus.The 528-bp open reading frame specified a primary translation product of 175 amino acids with a calculatedM r of 18,789. The predicted amino acid sequence of the DHQase showed similarities to bacterial and fungal type II DHQases. Overexpression of thedhqgene was accomplished inEscherichia coliusing a gene fusion technique in which amalE,the gene encoding the maltose binding protein (MBP), was fused via a short oligonucleotide region to the beginning ofdhq.The recombinant MBP-DHQase fusion protein was purified by affinity chromatography and cleaved using thrombin. The resulting DHQase, separated from the MBP, demonstrated typical properties of a type II DHQase: a relatively highK m for the dehydroquinate substrate (650 μM) and extreme thermal stability. The subunitM r estimated by SDS-PAGE was 19,000, and the nativeM r estimated by gel-exclusion chromatography and sucrose-density centrifugation was 130,000, suggesting that the enzyme is a homoheptamer (type II DHQases are typically homododecamers). The MBP-DHQase complex also adopted a heptameric structure and was a thermostable, fully active DHQase, indicating that the N-terminus is not involved in formation of protomer-protomer complexes. Previous analyses have supported positioning the N-terminus of type II DHQases close to the active site and a conformational change in this region coincident with ligand binding. Nonetheless, theK m and relativek c a t obtained for MBP-DHQase were indistinguishable from those observed for DHQase. Inactivation data of the DHQase fromS. hygroscopicuswith the arginine-specific reagent phenylglyoxal showed that a modified Arg residue(s) is likely close to the N-terminus and active site of DHQase, but does not play an essential role in catalysis.