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Barley limit dextrinase: Determinants of substrate specificity and sensitivity to the endogenous proteinaceous inhibitor
B. SVENSSON (1), S. Andersen (2), M. Møller (2), T. Olsen (2), S. Olesen (2), I. André (3) (1) Enzyme and Protein Chemistry, Dept Systems Biology, Technical University of Denmark, Lyngby, Denmark; (2) Enzyme and Protein Chemistry, Dept. Systems Biology, Technical University of Denmark, Lyngby, Denmark; (3) Dept. Biochem. Struct. Biol., Lund University, Lund, Sweden.

Cereal limit dextrinases (LDs) and endogenous limit dextrinase inhibitors (LDIs) are key players in degradation of a-1,6 glucosidic linkages from amylopectin and thus in starch mobilization of particular relevance in malting. Moreover LD and LDI are proposed to be important in starch biosynthesis. Notably, LD and LDI appeared essential in malting. Several crystal structures are determined of barley LD in complex with substrates, substrate analogues [1] and the potent endogenous LDI having picomolar affinity for LD [2]. Guided by the crystal structures we used site-directed mutagenesis to unravel determinants for substrate specificity and sensitivity to regulation of LD activity by LDI. Secondly, we examined the role of positions in the non-catalytic N-terminal domain of barley LD, which as reported by others may play a role in a variant of sorghum LD for elevated LD activity resulting in more digestible starch from this drought tolerant crop. Thus a finding connected with LD’s proposed role in starch biosynthesis [3]. In complementary studies computational design of LDI variants will be validated. The results are of interest besides to malting industries and cereal breeders, particularly to industries producing starch degrading enzymes and manufacture of starch products such as syrops. The findings are promising for future interdisciplinary achievements combining rational protein engineering with cereal crop improvements. This work is supported by a DTU PhD scholarship (SA) and a Research Talent grant from Danish Council for Independent Research | Technology and Production Sciences (MSM).