Lignocellulosic biomass bioconversion is hampered by the structural and chemical complexity of the network created by constituting polymers (cellulose, hemicellulose and lignin) that prevents reaching an effective yield in monomeric sugars (glucose, xylose) for subsequent conversion in biofuels for instance. Thus, a physicochemical pretreatment is an essential prerequisite to overcome these limitations and to render the biomass more accessible to enzymes.Hemicellulases have been repeatedly shown to improve enzymatic saccharification and can be used during cellulose conversion or in combination of chemical pretreatments. However the efficiency of hemicellulases can also be restricted owing to steric hindrance within the wall network and non-specific binding to lignin, thereby ending with limitations in enzymes penetration and progression within lignocelluloses. Combined microscopic techniques such as (immuno)-cytochemistry and micro-spectrophotometry provide insights into the structural change of cell walls after degradation and into enzyme distribution at the cell wall level. But these tools generally give static mapping data. Actually, data related to dynamics of enzymes during the degradation process in such complex substrates are essential to obtain.To gain this information, we have prepared some fluorescent polysaccharides and xylanase probes and their mobility was followed by measuring the fluorescence recovery after photobleaching in cell walls from poplar stems which have been pretreated differently using alkali, acid chlorite or hydrothermal treatments. The probes mobility reveals that, for non catalytic probes, progression is directly correlated to the loss or modification of the lignin while enzymatic probes progression is least in drastically treated substrates. Overall, probes progression is more influenced by their types and size than by pre-treatment type and cellular localization. This paves the way for investigating other enzymes and for correlating their progression with saccharification.

Tracking fluorescent enzymes to unravel limitations to lignocellulosic biomass degradation

247th National Meeting (2014)

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