Characterization of xyloglucan endotransglycosylase (XET) enzymes from different plant species

Kingdom Plantae, which has extensive variety between species and provides the planet with most of the fixed organic carbon, is a very important kingdom with a specific distinctive feature; cell walls made of polysaccharides and functional and structural proteins. Plant cell walls are made of cellulose microfibrils embedded in a matrix of pectic polysaccharides and cross-linked with non-cellulosic β-linked polysaccharides. Xyloglucan is a major non-cellulosic β-linked polysaccharide, it is ubiquitous in the cell walls of non-graminaceous, graminaceous and evolutionarily distant species like Equisetum plants, and it links cellulose microfibrils to form load-bearing structures in the primary cell wall. The cell wall changes during the lifespan of a plant. This change, or adaptation, is achieved by enzymes that are capable of modifying cell wall polysaccharides. Xyloglucan endotransglycosylase/hydrolase enzymes (XTH), found as multi-member isozyme families in most plant species, including agriculturally important species, catalyse endotransglycosylation and/or hydrolysis of xyloglucan and possibly other polysaccharides within the plant cell wall, resulting in modifications of the wall. XTH enzymes group into six phylogenetic clades depending on their amino acid sequences, which are also proven to differ in their substrate specificity. As demonstrated in this thesis, Group I XTH family members show dominant preference to xyloglucan whereas ancestral clade members have a broader substrate specificity, with Arabidopsis ATXTH3 members acting preferentially on mixed-linked β-glucan. On the other hand, with the discovery of mixed-linked β-glucan in Equisetum fluviatile, a new enzyme was identified named as mixed-linked β-glucan: xyloglucan endotransglycosylase (MXE). E. telmateia, an Equisetum species commonly found in Turkey, was used as an explant in order to purify, characterize and identify an MXE-like enzyme. With enzyme characterizations of MXE, ancestral clade and Group I members, evolution of XTHs in plants from bacterial licheninase ancestors is aimed to be enlightened.