amyP, a reporter gene to study strain degeneration in Clostridium acetobutylicum ATCC 824.

Clostridium acetobutylicum produces an extracellular alpha-amylase when grown on glucose as the sole carbon source. This enzyme was previously characterized from a biochemical point of view but its encoding gene was never identified. The 2283-bp amyP gene encodes a 83013-Da mature protein with an N-terminal domain that exhibits strong identity to the family 13 glycosyl hydrolases such as the Bacillus alpha-amylases. Transcriptional analysis revealed that amyP is transcribed in solventogenic but not in acidogenic chemostat cultures. These results are in agreement with the extracellular alpha-amylase activities indicating that the expression of amyP is regulated at the transcriptional level. amyP is located on the pSOL1 megaplasmid that carries all the genes involved in the final steps of solvent formation. Degeneration of C. acetobutylicum has been associated to the loss of pSOL1. We demonstrate here that amyP can be used as a reporter system to quantitatively follow this phenomenon.