The genes coding for d-ribulose-1,5-bisphosphate carboxylase (RuBPCase) from an iron-oxidizing bacterium, Thiobacillus ferrooxidans, were cloned into an Escherichia coli plasmid, pUC18. The recombinant plasmid, termed pTR11, contained a 4.0-kb PstI fragment including the entire coding regions for both large and small subunits of RuBPCase. Escherichia coli carrying pTR11 did not show any CO2-fixing activity. However, a derivative plasmid with an appropriate deletion, which was placed under the control of a tac promoter, conferred ribulose bisphosphate-dependent CO2-fixing activity on the host cell. Analysis of gel-filtration chromatography of the RuBPCase synthesized in E. coli revealed that it had a hexadecameric form like the native enzyme of T. ferrooxidans.