title: The debranching enzyme complex missing in glycogen accumulating mutants of Chlamydomonas reinhardtii displays an isoamylase-type specificity.
abstract: To investigate the functions of debranching enzymes in starch biosynthesis, we have partially purified and characterized these activities from wild type and mutant sta7 Chlamydomonas reinhardtii. Mutants of the STA7 locus substitute synthesis of insoluble granular starch by that of small amounts of glycogen-like material. The mutants were previously shown to lack an 88 kDa debranching enzyme. Two distinct debranching activities were detected in wild-type strains. The 88 kDa debranching enzyme subunit missing in glycogen-producing mutants (CIS1) is shown to be part of a multimeric enzyme complex. A monomeric 95 kDa debranching enzyme (CLD1) cleaved alpha-1,6 linkages separated by as few as three glucose residues while the multimeric complex was unable to do so. Both enzymes were able to debranch amylopectin while the alpha-1,6 linkages of glycogen were completely debranched by the multimeric complex only. Therefore CLD1 and the multimeric debranching enzyme display respectively the limit-dextrinase (pullulanase) and isoamylase-type specificities. Various mutations in the STA7 locus caused the loss of both CIS1 and of the multimeric isoamylase complex. In contrast to rice and maize mutants that accumulate phytoglycogen owing to mutation of an isoamylase-type DBE, isoamylase depletion in Chlamydomonas did not result in any qualitative or quantitative difference in pullulanase activity.
authors.affiliation: Laboratoire de Chimie Biologique, Unité Mixte de Recherche du C.N.R.S. no 8576, Bâtiment C9, Cité Scientifique, Université des Sciences et Technologies de Lille, 59655 Cedex, Villeneuve d'Ascq, France
authors.initials: D
authors.lastname: Dauvillée
authors.initials: V
authors.lastname: Mestre
authors.initials: C
authors.lastname: Colleoni
authors.initials: M
authors.lastname: Slomianny
authors.initials: G
authors.lastname: Mouille
authors.initials: B
authors.lastname: Delrue
authors.initials: C
authors.lastname: d'Hulst
authors.initials: C
authors.lastname: Bliard
authors.initials: J
authors.lastname: Nuzillard
authors.initials: S
authors.lastname: Ball