Journal Article
Applied and Environmental Microbiology, vol. 77, iss. 22, pp. 7933-7941, 2011
Authors
Jae San Ryu, Semarjit Shary, Carl J. Houtman, Ellen A. Panisko, Premsagar Korripally, Franz J. St. John, Casey Crooks, Matti Siika-aho, Jon K. Magnuson, Kenneth E. Hammel
Abstract
ABSTRACT
Brown rot basidiomycetes have an important ecological role in lignocellulose recycling and are notable for their rapid degradation of wood polymers via oxidative and hydrolytic mechanisms. However, most of these fungi apparently lack processive (
exo
-acting) cellulases, such as cellobiohydrolases, which are generally required for efficient cellulolysis. The recent sequencing of the
Postia placenta
genome now permits a proteomic approach to this longstanding conundrum. We grew
P. placenta
on solid aspen wood, extracted proteins from the biodegrading substrate, and analyzed tryptic digests by shotgun liquid chromatography-tandem mass spectrometry. Comparison of the data with the predicted
P. placenta
proteome revealed the presence of 34 likely glycoside hydrolases, but only four of these—two in glycoside hydrolase family 5, one in family 10, and one in family 12—have sequences that suggested possible activity on cellulose. We expressed these enzymes heterologously and determined that they all exhibited endoglucanase activity on phosphoric acid-swollen cellulose. They also slowly hydrolyzed filter paper, a more crystalline substrate, but the soluble/insoluble reducing sugar ratios they produced classify them as nonprocessive. Computer simulations indicated that these enzymes produced soluble/insoluble ratios on reduced phosphoric acid-swollen cellulose that were higher than expected for random hydrolysis, which suggests that they could possess limited
exo
activity, but they are at best 10-fold less processive than cellobiohydrolases. It appears likely that
P. placenta
employs a combination of oxidative mechanisms and
endo
-acting cellulases to degrade cellulose efficiently in the absence of a significant processive component.
