In this paper development of a cell-free enzymatic cascade is described for conversion of brown seaweed carbohydrates (alginate and glucose from laminarin) to 2-keto-3-deoxygluconate (KDG), a precursor in bioplastics production. The process involves five enzymatic steps in two parallel pathways (alginate to KDG and glucose to KDG) including alginate depolymerization, cofactor recycling (NADH\/NAD+), and conversion of intermediates to KDG, all integrated in a one-pot reaction system. Conversion of alginate to KDG was achieved in a three-step enzymatic process, starting with depolymerization of alginate by one endo- (RmAly3) and one exo-acting alginate lyase (RmAly4), followed by reduction of spontaneously formed 4-deoxy-L-erythro-5-hexoseulose uronate (DEH) to KDG by a novel reductase (Adh51 from Rhodothermus marinus<\/em>). Cofactor recycling and conversion of glucose to gluconate were integrated by introduction of a novel glucose dehydrogenase (GDH16 from Thermobaculum<\/em> strain MAT<\/a>1704[E1]<\/a>\u00a0) to oxidize glucose to gluconate while simultaneously regenerating NADH from NAD+<\/sup>. Generated gluconate was converted to KDG via dehydration catalyzed by the dihydroxyacid dehydratase DhadSs. The KDG productivity of the enzymatic cascade was 1 g\/Lh from alginate and glucose, supplying a proof of concept of the cell free system for KDG production, but with DhadSs identified as the rate-limiting factor, resulting in conversion of 35% of the combined alginate and glucose addition to the system.<\/p>\n","protected":false},"excerpt":{"rendered":" Abstract In this paper development of a cell-free enzymatic cascade is described for conversion of brown seaweed carbohydrates (alginate and glucose from laminarin) to 2-keto-3-deoxygluconate (KDG), a precursor in bioplastics production. The process involves five enzymatic steps in two parallel pathways (alginate to KDG and glucose to KDG) including alginate depolymerization, cofactor recycling (NADH\/NAD+), and … Read more<\/a><\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"closed","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[1],"tags":[6,3,5,4],"class_list":["post-1","post","type-post","status-publish","format-standard","hentry","category-uncategorized","tag-2-keto-3-deoxygluconate","tag-alginate","tag-glucose-dehydrogenase","tag-reductase"],"_links":{"self":[{"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/posts\/1","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/seabiocat.net\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=1"}],"version-history":[{"count":3,"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/posts\/1\/revisions"}],"predecessor-version":[{"id":76,"href":"https:\/\/seabiocat.net\/index.php?rest_route=\/wp\/v2\/posts\/1\/revisions\/76"}],"wp:attachment":[{"href":"https:\/\/seabiocat.net\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=1"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/seabiocat.net\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=1"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/seabiocat.net\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=1"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}