Chitin hydrolysis and N-acetylglucosamine utilization by solventogenic clostridia

Abstract

Environment pollution and energy supply are among the huge problems which threaten the world, especially in industrialised countries. Several studies have considered how to exploit waste materials as renewable substrates for various industries to obtain different products. Some wastes from the aquatic food industry contain a considerable amount of the N-acetylglucosamine (NAG) polymer chitin, which has potential as a substrate for the solventogenic clostridia in the acetone-butanol-ethanol fermentation. Development of an effective process will, however, depend on a detailed understanding of the mechanism and control of chitin hydrolysis and NAG metabolism. Clostridium beijerinckii NCIMB 8052 was shown to exhibit chitinase activity and to be able to grown on NAG. The predominant mechanism for uptake of sugars and sugar derivatives in the clostridia is the phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS). Extracts of C.beijerinckii grown on NAG exhibited a phosphotransferase activity for NAG which was also present in extracts of cells grown on glucose, consistent with the observation that glucose did not repress utilization of NAG in media containing both substrates. Genomic analysis has identified genes encoding a putative nag-pts that belongs to the glucose family of PTS permeases. Two divergent genes encode the IIA and IICB domains of the PTS, and are associated with a gene encoding a putative transcriptional antiterminator. These genes were found to be expressed in cells growing on NAG or glucose, but not glucitol. The role of the putative nag-pts genes in NAG uptake was confirmed by functional analysis. An artificial NAG operon was constructed in which the nag-pts genes were in series and expression of the operon in Escherichia coli mutants provided evidence for the ability of the PTS to transport and phosphorylate NAG and glucose, but not mannose.