Cloning and sequence analysis of the genes encoding phosphotransbutyrylase and butyrate kinase from Clostridium acetobutylicum NCIMB 8052

Abstract

An 8.1-kb fragment of chromosomal DNA from Clostridium acetobutylicum NCIMB 8052 (formerly NCIB 8052) has been cloned into plasmid pAT153 and shown to allow the growth of Escherichia coli LJ32 (F+ atoC2c atoD32 fadR) on butyrate as the sole source of carbon and energy. Deletion analysis delineated a 3.9-kb subfragment capable of complementation. The nucleotide sequence of this fragment was determined and it was shown to encode three complete, and two incomplete open reading frames (ORFs). Based on enzymic studies of recombinant clones, two of these ORFs were shown to encode phosphotransbutyrylase and butyrate kinase. The above enzymes are involved in the acidogenic phase of fermentation in C. acetobutylicum. The fragment also carries an incomplete ORF encoding a polypeptide exhibiting substantial similarity to dihydropteroate synthase.