Characterization Of Pectin Features That Distinctively Affect Lignocellulose Enzymatic Saccharification In Plants

Lignocellulosic ethanol has been defined as an excellent additive to gasoline with less net carbon release to the environment.In principle,lignocellulose ethanol process involves in three major steps: initial physical and chemical pretreatment for wall polymer depolymerization,sequential enzymatic hydrolysis for soluble sugar release and final yeast fermentation for ethanol production.However,as lignocellulose recalcitrance fundamentally decides an inacceptable costly biomass process,genetic modification of plant cell walls has been posed as a promising solution.Hence,it becomes important to sort out the mechanism of how plant cell walls distinctively affect biomass enzymatic saccharification under various chemical pretreatments,providing insights into plant cell wall biosynthesis and bioenergy crop breeding.Pectin is a major component of primary cell walls with minor deposition in the secondary cell walls.However,it has been characterized that pectin dynamically affects secondary wall biosynthesis and deposition.Although three major wall polymers(cellulose,hemicellulose,and lignin)have been reported about their distinct impacts on biomass enzymatic saccharification in different plant species,little is known about role of pectin in lignocellulose enzymatic hydrolysis process under various chemical pretreatments.Miscanthus has been considered as a leading bioenergy crop,due to its high biomass yield,well adaption to environmental conditions and diverse germplasm accessions.By comparison,Arabidopsis thaliana is a genetic model plant and its cell wall biosynthesis has been well known.In this study,we collected large populations of Miscanthus accessions and examined their pectin impacts on biomass enzymatic saccharification under various chemical pretreatments.Meanwhile,we characterized two Arabidopsis pectin methylesterase 1 gene(AtPME1;AtPME26)and investigated their functions in pectin modification.Finally,this study attempted to explore the mechanism of how pectin positively affects biomass enzymatic digestibility in plants.The major findings were described below.Using total 179 samples of representative Miscanthus accessions,we determined diverse pectin levels and largely varied hexoses yields released from enzymatic hydrolysis after chemical pretreatments.Integrative analysis indicated that pectin levels positively affect hexoses yields under acid or alkali pretreatments.Using four typical pairs of Miscanthus samples,this study found out that AO-extractable uronic acids,other than hexoses and pentoses,could significantly reduce lignocellulose crystallinity for largely enhanced biomass enzymatic saccharification,suggesting that genetic increasing of uronic acids may be an applicable approach for bioenergy Miscanthus breeding.Meanwhile,we explored PME1 and PME26 genes roles in pectin post-modification in Arabidopsis.As a result,the degree of methylesterification(DM)of galacturonic acid(GalA)was significantly increased in both pme1 and pme26 mutants,but their total GalA contents were little changed compared to the wild type.Further analysis indicated that other wall polymer features were altered in the mutants,leading to much more polymers extractable in vitro.Hence,the mutants showed much reduced lignocellulose crystallinity for hexoses yields increased by 28%-62% compared with the wild type.Finally,to further elucidate pectin roles in biomass enzymatic hydrolysis,this study used atomic force microscopy to in vitro observe pectin image in Arabidopsis,which could be applied to explore potential pectin interaction with other wall polymers in the future.