Mechanisms of laccase-mediator treatments improving the enzymatic hydrolysis of pre-treated spruce

Abstract

BackgroundThe recalcitrance of softwood to enzymatic hydrolysis is one of the major bottlenecks hindering its profitable use as a raw material for platform sugars. In softwood, the guaiacyl-type lignin is especially problematic, since it is known to bind hydrolytic enzymes non-specifically, rendering them inactive towards cellulose. One approach to improve hydrolysis yields is the modification of lignin and of cellulose structures by laccase-mediator treatments (LMTs).ResultsLMTs were studied to improve the hydrolysis of steam pre-treated spruce (SPS). Three mediators with three distinct reaction mechanisms (ABTS, HBT, and TEMPO) and one natural mediator (AS, that is, acetosyringone) were tested. Of the studied LMTs, laccase-ABTS treatment improved the degree of hydrolysis by 54%, while acetosyringone and TEMPO increased the hydrolysis yield by 49% and 36%, respectively. On the other hand, laccase-HBT treatment improved the degree of hydrolysis only by 22%, which was in the same order of magnitude as the increase induced by laccase treatment without added mediators (19%). The improvements were due to lignin modification that led to reduced adsorption of endoglucanase Cel5A and cellobiohydrolase Cel7A on lignin. TEMPO was the only mediator that modified cellulose structure by oxidizing hydroxyls at the C6 position to carbonyls and partially further to carboxyls. Oxidation of the reducing end C1 carbonyls was also observed. In contrast to lignin modification, oxidation of cellulose impaired enzymatic hydrolysis.ConclusionsLMTs, in general, improved the enzymatic hydrolysis of SPS. The mechanism of the improvement was shown to be based on reduced adsorption of the main cellulases on SPS lignin rather than cellulose oxidation. In fact, at higher mediator concentrations the advantage of lignin modification in enzymatic saccharification was overcome by the negative effect of cellulose oxidation. For future applications, it would be beneficial to be able to understand and modify the binding properties of lignin in order to decrease unspecific enzyme binding and thus to increase the mobility, action, and recyclability of the hydrolytic enzymes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-014-0177-8) contains supplementary material, which is available to authorized users.

Laccases as a Potential Tool for the Efficient Conversion of Lignocellulosic Biomass: A Review

“…Consequently, an enhancement of saccharification yield by 13% was observed during the subsequent enzymatic hydrolysis. Similar results were attained by Moilanen et al [117,118] when acid steam-pretreated spruce was treated with C. unicolor and T. hirsuta laccases. However, using acid steam-pretreated giant read (Arundo donax), C. unicolor laccase reduced the hydrolysis yield by 17% [117].…”

“…These results were comparable to those obtained with P. cinnabarinus laccase-HBT as LMS. Moilanen et al [118], in contrast, observed an increase in the lignin content of acid-steam pretreated spruce when using acetosyringone mediator together with T. hirsuta laccase. In spite of this effect, laccase-acetosyringone treatment improved the hydrolysis yield by 36%.…”

“…13% increase in sugar yield [116] Acid steam-pretreated spruce Cerrena unicolor and T. hirsuta laccases Reduced binding of hydrolytic enzymes by lignin modification Improvement of hydrolysis yield by 12% [117,118] Steam-exploded sugarcane bagasse G. lucidum Delignification 75% increase in glucose production [119] Corncob residue…”

“…By using ABTS and TEMPO as mediators of T. hirsuta laccase, Moilanen et al [118] increased the hydrolysis yields of acid steam-exploded spruce by 54% and 49%, respectively. These improvements were explained to be based on the reduction of the unspecific adsorption of hydrolases on enzyme-treated lignin.…”

Bioprocessing of wheat bran for the production of lignocellulolytic enzyme cocktail byCotylidia pannosaunder submerged conditions

“…9,10 For biological pre-treatment, laccase producing brown rot and white rot basidiomycete fungal strains have been identified as the best delignifying organisms. 11 A number of white rot fungi are reported for the biological pretreatment or simultaneous saccharification and fermentation of lignocellulosic biomass.…”