title: Glucan, water dikinase phosphorylates crystalline maltodextrins and thereby initiates solubilization.
abstract: Starch phosphorylation by glucan, water dikinase (GWD; EC 2.7.9.4) is an essential step in the breakdown of native starch particles, but the underlying mechanisms have remained obscure. In this paper, the initial reactions of starch degradation were analyzed using crystallized maltodextrins as model carbohydrates. As revealed by X-ray diffraction analysis, the crystallized maltodextrins represent the B-type starch allomorph. Recombinant GWD phosphorylated crystalline maltodextrins with a high specific activity (55-60 nmol mg-1 protein min-1), but exhibited very little activity with the same maltodextrins that had been solubilized by heat treatment. Recombinant phosphoglucan, water dikinase (PWD; EC 2.7.9.5) utilized the crystalline maltodextrins only when pre-phosphorylated by GWD. Phosphorylation of crystalline maltodextrins, as catalyzed by GWD, initiated solubilization of neutral as well as phosphorylated glucans. In both the insoluble and the soluble state, mono-, di- and triphosphorylated alpha-glucans were observed, with wide and overlapping ranges of degree of polymerization. Thus, the substrate specificity of the GWD is defined by the physical arrangement of alpha-glucans rather than by structural parameters, such as the distribution of branching points or degree of polymerization. Unlike GWD and PWD, recombinant beta-amylase isozyme 3 (BAM3), which has been shown to be essential for plastidial starch degradation, preferentially degraded soluble maltodextrins rather than crystallized glucans. In summary, two conclusions were reached. Firstly, carbohydrate targets of GWD are primarily defined by the molecular order of glucan helices. Secondly, GWD-catalyzed phosphorylation mediates the phase transition of glucans from a highly ordered to a less ordered and hydrated state.
authors.affiliation: UP Transfer GmbH, Am Neuen Palais 10, Building 6, 14469 Potsdam, Germany.
authors.forename: Mahdi
authors.initials: M
authors.lastname: Hejazi
authors.forename: Joerg
authors.initials: J
authors.lastname: Fettke
authors.forename: Sophie
authors.initials: S
authors.lastname: Haebel
authors.forename: Christoph
authors.initials: C
authors.lastname: Edner
authors.forename: Oskar
authors.initials: O
authors.lastname: Paris
authors.forename: Claus
authors.initials: C
authors.lastname: Frohberg
authors.forename: Martin
authors.initials: M
authors.lastname: Steup
authors.forename: Gerhard
authors.initials: G
authors.lastname: Ritte
mesh_terms: D010766:Phosphorylation; D017866:Phosphotransferases (Paired Acceptors) / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D010944:Plants / Q000201:enzymology; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility; D013213:Starch / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D010940:Plant Proteins; D011134:Polysaccharides; C008315:maltodextrin; D013213:Starch; D017866:Phosphotransferases (Paired Acceptors)