Medium chain length polyhydroxyalkanoate produced from ethanol by Pseudomonas putida grown in liquid obtained from acidogenic digestion of organic municipal solid waste.

Production of medium chain length polyhydroxyalkanoate (mcl-PHA) up to about 6 g.L-1 was obtained by feeding ethanol to Pseudomonas putida growing in liquid obtained from acidogenic digestion of organic municipal solid waste. Washing the wet, heat-inactivated Pseudomonas cells at the end of the fermentation with ethanol obviated the need of drying the biomass and enabled the removal of contaminating lipids before solvent-mediated extraction of PHA. Using 'green' solvents, 90 to near 100% of the mcl-PHA was extracted and purities of 71-78% mcl-PHA were reached already by centrifugation and decantation without further filtration for biomass removal. The mcl-PHA produced in this way consists of 10-18% C8, 72-78% C10 and 8-12% C12 chains (entirely medium chain length), has a crystallinity and melting temperature of ∼13% and ∼49 °C, respectively, and is a stiff rubberlike, colourless material at room temperature.

Characterization of the starvation-induced chitinase CfcA and α-1,3-glucanase AgnB of Aspergillus niger.

The common saprophyte Aspergillus niger may experience carbon starvation in nature as well as during industrial fermentations. Starvation survival strategies, such as conidiation or the formation of exploratory hyphae, require energy and building blocks, which may be supplied by autolysis. Glycoside hydrolases are key effectors of autolytic degradation of fungal cell walls, but knowledge on their identity and functionality is still limited. We recently identified agnB and cfcA as two genes encoding carbohydrate-active enzymes that had notably increased transcription during carbon starvation in A. niger. Here, we report the biochemical and functional characterization of these enzymes. AgnB is an α-1,3-glucanase that releases glucose from α-1,3-glucan substrates with a minimum degree of polymerization of 4. CfcA is a chitinase that releases dimers from the nonreducing end of chitin. These enzymes thus attack polymers that are found in the fungal cell wall and may have a role in autolytic fungal cell wall degradation in A. niger. Indeed, cell wall degradation during carbon starvation was reduced in the double deletion mutant ΔcfcA ΔagnB compared to the wild-type strain. Furthermore, the cell walls of the carbon-starved mycelium of the mutant contained a higher fraction of chitin or chitosan. The function of at least one of these enzymes, CfcA, therefore appears to be in the recycling of cell wall carbohydrates under carbon limiting conditions. CfcA thus may be a candidate effector for on demand cell lysis, which could be employed in industrial processes for recovery of intracellular products.