Ulvan lyase from Formosa agariphila and its applicability in depolymerisation of ulvan extracted from three different Ulva species

Artikel i vetenskaplig tidskrift, 2018

Members of green macroalgae cause green tides in eutrophicated coastal waters. These green tides pose an environmental issue and an economic burden on coastal municipalities. The biomass from these green tides has a potential to be used as feedstock in biorefinery due to the content of interesting biomacromolecules. Ulvan, an anionic water-soluble polysaccharide, is one of such components, and its depolymerisation to high-value oligosaccharides or fermentable monosaccharides would bring value to green tide biomass which is otherwise left to decompose. However, only a few ulvan depolymerising enzymes are studied to date. Ulvan lyases depolymerise ulvan, via the β-elimination mechanism, leading to release of oligosaccharides with an unsaturated 4‑deoxy‑l‑threo‑hex‑4‑enopyranosiduronic acid at the non-reducing end. In this study, we have identified the presence of two different domains, a catalytic and a non-catalytic, in a putative ulvan lyase from Formosa agariphila KMM 3901. We overexpressed, purified, and biochemically characterised the full-length ulvan lyase, which was found to be most active at a temperature of 45 °C and pH 8.5. It exhibited high specificity for ulvan and did not degrade heparan sulphate, chondroitin sulphate, alginate, pectin or xanthan. Detailed analyses of end products of the enzymatic degradation of ulvan using 1H NMR and LC-MS revealed a disaccharide with an unsaturated uronic acid (∆) linked to 3‑sulphated rhamnose (Rha3S), trisaccharide with xylose (Xyl) flanked by Rha3S (Rha3S-Xyl-Rha3S), tetrasaccharide with an unsaturated uronic acid at the non-reducing end (∆Rha3S-Xyl-Rha3S) and pentasaccharides (Rha3S-Xyl-Rha3S-Xyl-Rha3S and branched ∆Rha3S-Xyl-(∆)Rha3S) as the principal end products. We also found that the catalytic domain that lacks the non-catalytic carbohydrate binding module exhibited higher affinity for the soluble ulvan and efficiently depolymerised it. This study reveals the characteristics of the endolytic ulvan lyase, which is a member of the ulvan utilisation loci in Formosa, and points towards the potential ulvan depolymerisation applications in Ulva biorefinery.