Characterization of an acetyl-CoA C-acetyltransferase (thiolase) gene from Clostridium acetobutylicum ATCC 824.

Thiolase (Thl) is an important enzyme at the junction in the pathway leading to the production of either acids (acetate or butyrate) or solvents (acetone, butanol or ethanol) during the growth of Clostridium acetobutylicum ATCC 824. Cloning and expression of the Thl-encoding gene (thl) has been described [Petersen and Bennett, Appl. Environ. Microbiol. 57 (1991) 2735-2741], as has the purification and properties of the enzyme [Wiesenborn et al., Appl. Environ. Microbiol. 54 (1988) 2717-2722]. Here, we present the complete nucleotide sequence (1.9 kb) of thl. The gene encodes a protein of 392 amino acids (aa) (41,237 Da), which mass is in agreement with previous findings using the purified protein. Primer extension analysis has defined the promoter region, and a stem-loop structure found at the end of thl indicates that it is not part of an operon. The aa sequence of Thl showed homology to those of four other beta-ketothiolases: (i) PhbC of Alcaligenes eutrophus, (ii) PhbA of Chromatium vinosum, (iii) PhbA of Thiocystis violacea and (iv) PhbA of Zoogloea ramigera. The C terminus of an open reading frame found upstream from the Thl sequence is similar to OrfX of Bacillus subtilis and to NfrC of Escherichia coli.