Mass Spectrometry Deduces Selectivity of Glycoside Hydrolases for Degrading Biomass Polysaccharides

Improving the annotation of glycoside hydrolases and their phylogenetic trees.

The Science

Multiple classes of polysaccharide-degrading enzymes are used to hydrolyze plant biomass into fermentable sugars for conversion to biofuels. However, there are large numbers of suspected polysaccharide-degrading enzymes whose activities have not been determined biochemically. Researchers have now determined the reaction specificity and other parameters for several of these uncharacterized enzymes using a special mass spectroscopy system along with artificial substrates.

The Impact

Improving the annotation of glycoside hydrolase (GH) phylogenetic trees will improve understanding of the function, synergy, and stability of these enzymes and thereby the creation of biomass-degrading enzymatic cocktails.


Researchers at the Department of Energy’s (DOE) Great Lakes Bioenergy Research Center (GLBRC) have used chemically synthesized nanostructure-initiator mass spectrometry (NIMS) probes derivatized with tetrasaccharides to study the reactivity of several enzymes representative of GH function. Patterns of reactivity identified with these NIMS probes provide a diagnostic approach to assess reaction selectivity as well as comparative apparent rate information. Their results show diagnostic patterns for reactions of a β-glucosidase, relaxed but varied specificity of several endoglucanases, and high specificity of a cellobiohydrolase with the model substrate. The researchers also modeled time-dependent reactions of these enzymes by numerical integration, providing a quantitative basis to make functional distinctions among reactive properties, thus providing a new approach to enhance the annotation of GH phylogenetic trees with functional measurements. This research was carried out in collaboration with researchers at DOE’s Joint BioEnergy Institute (JBEI).

Principal Investigator(s)

Brian Fox
University of Wisconsin-Madison


GLBRC and JBEI are supported by DOE’s Office of Science, Office of Biological and Environmental Research through contracts DE-FC02-07ER64494 and DE-AC02-05CH11231, respectively.


Deng, K., T. E. Takasuka, C. M. Bianchetti, L. F. Bergeman, P. D. Adams, T. R. Northen, and B. G. Fox. 2015. “Use of Nanostructure-Initiator Mass Spectrometry to Deduce Selectivity of Reaction in Glycoside Hydrolases,” Frontiers in Bioengineering and Biotechnology 3(165), DOI: 10.3389/fbioe.2015.00165.