Ulvan, which is one of the major structural polysaccharides of the cell walls of green macroalgae, is degraded by ulvan lyases via the β-elimination mechanism with the release of oligosaccharides that have unsaturated 4-deoxy-L-threo-hex-4-enopyranosiduronic acid (∆) at the non-reducing end. These ulvan lyases belong to the PL24 or PL25 or PL28 family in the CAZy database. In this study, we identify and biochemically characterise a periplasmic novel broad-spectrum ulvan lyase from Formosa agariphila KMM 3901. The lyase was overexpressed in Escherichia coli, and the purified recombinant enzyme depolymerised ulvan in an endolytic manner with a Km of 0.77 mg/ml, and displayed optimum activity at 40 °C and pH 8. This lyase also degraded heparan sulphate and chondroitin sulphate. Detailed analyses of the end-products of the enzymatic degradation of ulvan using 1H- and 13C-NMR and LC-MS revealed an unsaturated disaccharide (∆Rha3S) and a tetrasaccharide (∆Rha3S-Xyl-Rha) as the principal end-products. In contrast to the previously described ulvan lyases, this novel lyase is mostly composed of α-helices that form an (α/α)6 incomplete toroid domain and displays a remarkably broad-spectrum activity. This novel lyase is the first member of a new family of ulvan lyases.

Globally, there is a need for novel vegetarian protein sources. We recently showed that the pH-shift process, using alkaline protein solubilization followed by isoelectric precipitation, is an efficient way to produce extracts with high protein concentrations from Ulva lactuca (>50% on a dry matter basis). However, the total protein yield was low, and to improve this, the effects of adding ulvan lyase, preincubating the seaweed homogenate at pH 8.5 and using different protein extraction temperatures (8 °C, RT and 40 °C), were evaluated in this study. Addition of ulvan lyase reduced protein solubility but increased the precipitation. Incubation at pH 8.5, without ulvan lyase added, significantly increased both protein solubility and precipitation at 8 °C and RT. Temperature per se had no effect on protein solubility, while protein precipitation increased with decreasing temperature. Highest protein yield (29%) was achieved when keeping the process at 8 °C with a preincubation step at pH 8.5 for 1 h. By these process modifications, the yield was 3.2 times higher than achieved by the control process (9.2%).

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