Engineered yeast whole-cell biocatalyst for direct degradation  of alginate from macroalgae and production  of non-commercialized useful monosaccharide from alginate.
Alginate is a major component of brown macroalgae. In macroalgae, an endolytic alginate lyase first degrades alginate into oligosaccharides. These oligosaccharides are further broken down into monosaccharides by an exolytic alginate lyase. In this study, genes encoding various alginate lyases derived from alginate-assimilating marine bacterium Saccharophagus degradans were isolated, and their enzymes were displayed using the yeast cell surface display system. Alg7A-, Alg7D-, and Alg18J-displaying yeasts showed endolytic alginate lyase activity. On the other hand, Alg7K-displaying yeast showed exolytic alginate lyase activity. Alg7A, Alg7D, Alg7K, and Alg18J, when displayed on yeast cell surface, demonstrated both polyguluronate lyase and polymannuronate lyase activities. Additionally, polyguluronic acid could be much easily degraded by Alg7A, Alg7K, and Alg7D than polymannuronic acid. In contrast, polymannuronic acid could be much easily degraded by Alg18J than polyguluronic acid. We further constructed yeasts co-displaying endolytic and exolytic alginate lyases. Degradation efficiency by the co-displaying yeasts were significantly higher than single alginate lyase-displaying yeasts. Alg7A/Alg7K co-displaying yeast had maximum alginate degrading activity, with production of 1.98 g/L of reducing sugars in a 60-min reaction. This system developed, along with our findings, will contribute to the efficient utilization and production of useful and non-commercialized monosaccharides from alginate by Saccharomyces cerevisiae.
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan.|JST, CREST, Kawaguchi, Saitama, Japan.|Japan Society for the Promotion of Science, Sakyo-ku, Kyoto, Japan.
Toshiyuki
T
Takagi
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan.|JST, CREST, Kawaguchi, Saitama, Japan.
Takahiro
T
Yokoi
JST, CREST, Kawaguchi, Saitama, Japan.|Department of Life Sciences, Graduate School of Bioresources, Mie University, Tsu, Mie, Japan.
Toshiyuki
T
Shibata
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan.|JST, CREST, Kawaguchi, Saitama, Japan.
Hironobu
H
Morisaka
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan.|JST, CREST, Kawaguchi, Saitama, Japan.
Kouichi
K
Kuroda
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, Japan. miueda@kais.kyoto-u.ac.jp.|JST, CREST, Kawaguchi, Saitama, Japan. miueda@kais.kyoto-u.ac.jp.
Mitsuyoshi
M
Ueda
D000464:Alginates / Q000378:metabolism*; D020723:Glucuronic Acid / Q000378:metabolism; D006603:Hexuronic Acids / Q000378:metabolism; D006868:Hydrolysis; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009005:Monosaccharides / Q000378:metabolism*; D011133:Polysaccharide-Lyases / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D012624:Seaweed / Q000378:metabolism*
D000464:Alginates; D006603:Hexuronic Acids; D008565:Membrane Proteins; D009005:Monosaccharides; D020723:Glucuronic Acid; D011133:Polysaccharide-Lyases; C016158:poly(beta-D-mannuronate) lyase