Recombinant alginate lyases and mannitol dehydrogenase enhance hydrolysis of macroalgal carbohydrates

Brown macroalgae are a promising source for bioethanol production, primarily due to their high carbohydrate, low lignin and high moisture content. Bioconversion of macroalgae to ethanol requires a yeast, such as Saccharomyces cerevisiae, that can hydrolyse the macroalgal carbohydrates, namely laminarin, mannitol and alginate. In this study, the mannitol dehydrogenase (MDH) genes from Aspergillus fumigatus (AfMDH) and Talaromyces islandicus (TiMDH), and the alginate lyase (AL) genes from Sphingomonas sp. (SpxAL and SpeAL) and Talaromyces emersonii (TeeAL) were expressed in the laboratory strain, S. cerevisiae Y294. Co-cultures of a laminarinase-producing yeast, Y294[Relam1/Tvlam1] and yeasts expressing mannitol dehydrogenases and alginate lyases were evaluated for the consolidated bioprocessing of the major carbohydrates in brown macroalgae. Laminarin and mannitol were targeted for ethanol production, while alginate was depolymerised to expose mannitol. A co-culture of S. cerevisiae Y294[Relam1/Tvlam1], [AfMDH] and [TeeAL/SpxAL] strains produced 10.30 g/L ethanol from Ecklonia maxima, representing a 98 % carbon conversion (based on the laminarin and mannitol content). A strain expressing both endo- and exo-alginate lyase improved the ethanol yield by 42.28 % compared to strains expressing only laminarinase- and mannitol dehydrogenase. Scanning electron microscopy further revealed that co-cultures containing laminarinase, MDH, and AL enzymes promoted significant physical degradation and increased porosity in macroalgal substrates, suggesting enhanced alginate hydrolysis and improved enzyme accessibility. This is the first report on the simultaneous hydrolysis of mannitol, alginate and laminarin with recombinant enzymes during macroalgal fermentation. The results demonstrate significant progress towards exploiting brown macroalgae for bioconversion to ethanol and high-value products.