Monday, May 2, 2011
Grand Ballroom C-D, 2nd fl (Sheraton Seattle)
Caldicellulosiruptor bescii and Caldicellulosiruptor obsidiansis are extremely thermophilic, Gram-positive bacteria that efficiently degrade both cellulose and hemicellulose, making them relevant model systems for the deconstruction of lignocellulosic biomass for biofuel production. In order to investigate the cellulolytic protein apparatus of these two closely related species, label-free quantitative proteomics was used to compare protein abundance in their secretomes over the course of crystalline cellulose fermentation. Both organisms’ secretomes consisted of over 400 proteins, the most abundant being multidomain glycosidases, extracellular solute binding proteins, flagellin, putative pectate lyases, and uncharacterized proteins with predicted secretion signals. Mass spectrometric characterizations together with cellulase activity measurements revealed a substantial abundance increase of some bifunctional multidomain glycosidases composed of glycosidase (GH) domain families 5, 9, 10, 44 or 48 and family 3 carbohydrate binding modules (CBM3). In addition to their orthologous cellulases, the organisms expressed unique glycosidases distinguished by different domain organization. Additionally, mass spectrometric data for late exponential stage C. obsidiansis cultures across different carbon sources revealed significantly higher glycosidase abundance on cellobiose than on the crystalline cellulose substrates Avicel and filter paper. The cell lysates contained over 1400 proteins, including additional GH domain family 3, 4 and 31 enzymes as well as a putatively membrane bound GH5. When C. obsidiansis was grown on switchgrass, the abundance of hemicellulases and certain sugar transporters increased significantly compared to the other substrates. Cultivation on biomass also caused an 80-fold increase of a xylose isomerase, a protein that has been shown to facilitate ethanol production from hemicellulose.