By replacing a native promoter with lac and tac promoters, the gene encoding an ethylene-forming enzyme (EFE) from Pseudomonas syringae pv. phaseolicola PK2 was overexpressed in Escherichia coli. The EFE protein expressed by a multicopy plasmid accounted for more than 30% of the total cellular protein, resulting in ethylene-forming activities higher than 10 mu l of ethylene (mg cell)(-1)h(-1) in recombinant E. coli cells. However, most of the EFE protein accumulated as inactive inclusion bodies particularly at elevated temperatures (>30 degrees C). We present an efficient procedure for reconstituting an active enzyme from inclusion bodies by solubilization with 8 M urea and dialysis. The reconstituted EFE has specific activity identical to that of the native enzyme from P. syringae, suggesting that the EFE protein has an intrinsic folding capability in vitro.