Characterization of Carbohydrate Accessibility and Enzyme Adsorption Capacity for Diverse Cell Wall Phenotypes Subjected to Alkaline Hydrogen Peroxide Pretreatment

This study characterizes the changes to polysaccharides in terms of their abundance, accessibility, and affinity for binding by a range of cellulolytic enzymes and CBMs in the cell walls of grasses and dicots that were alkaline hydrogen peroxide (AHP) pretreated at varied severities or hydrothermally pretreated followed by AHP post-treatment.  These bioenergy feedstocks represent relatively diverse cell wall phenotypes, including the monocots switchgrass (Panicum virgatum cv. Cave-In-Rock), corn stovers (a commercial hybrid and inbred brown midrib lines bm1 and bm3), and Miscanthus spp. and herbaceous and woody dicots including goldenrod (Solidago spp.) and hybrid poplar (Populus spp.).  It was shown that increasing H₂O₂ loading results in increasing digestibility only for monocot grasses, while the digestibility of dicots is saturated at low H₂O₂ loadings. Polysaccharide accessibility in the residual cell wall is characterized through “glycome profile” which involves screening successive cell wall extracts with an array of 156 monoclonal antibodies with affinities for the entire spectrum of non-cellulosic cell wall polysaccharides revealing changes in their accessibility and extractability as a function of pretreatment.   Enzyme binding is assessed by characterizing binding isotherms for the proteins contained commercial cellulase enzyme cocktails, purified cellulase components, and a library of GFP-linked CBMs.