title: In vitro Biochemical Characterization of All Barley Endosperm Starch Synthases.
abstract: Starch is the main storage polysaccharide in cereals and the major source of calories in the human diet. It is synthesized by a panel of enzymes including five classes of starch synthases (SSs). While the overall starch synthase (SS) reaction is known, the functional differences between the five SS classes are poorly understood. Much of our knowledge comes from analyzing mutant plants with altered SS activities, but the resulting data are often difficult to interpret as a result of pleitropic effects, competition between enzymes, overlaps in enzyme activity and disruption of multi-enzyme complexes. Here we provide a detailed biochemical study of the activity of all five classes of SSs in barley endosperm. Each enzyme was produced recombinantly in E. coli and the properties and modes of action in vitro were studied in isolation from other SSs and other substrate modifying activities. Our results define the mode of action of each SS class in unprecedented detail; we analyze their substrate selection, temperature dependence and stability, substrate affinity and temporal abundance during barley development. Our results are at variance with some generally accepted ideas about starch biosynthesis and might lead to the reinterpretation of results obtained in planta. In particular, they indicate that granule bound SS is capable of processive action even in the absence of a starch matrix, that SSI has no elongation limit, and that SSIV, believed to be critical for the initiation of starch granules, has maltoligosaccharides and not polysaccharides as its preferred substrates.
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Jose A
authors: JA
authors: Cuesta-Seijo
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Morten M
authors: MM
authors: Nielsen
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Christian
authors: C
authors: Ruzanski
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Katarzyna
authors: K
authors: Krucewicz
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Sophie R
authors: SR
authors: Beeren
authors: Department of Plant and Environmental Sciences, University of Copenhagen Frederiksberg, Copenhagen, Denmark.
authors: Maja G
authors: MG
authors: Rydhal
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Yayoi
authors: Y
authors: Yoshimura
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Alexander
authors: A
authors: Striebeck
authors: Department of Plant and Environmental Sciences, University of Copenhagen Frederiksberg, Copenhagen, Denmark.
authors: Mohammed S
authors: MS
authors: Motawia
authors: Department of Plant and Environmental Sciences, University of Copenhagen Frederiksberg, Copenhagen, Denmark.
authors: William G T
authors: WG
authors: Willats
authors: Carlsberg Research Laboratory Copenhagen, Denmark.
authors: Monica M
authors: MM
authors: Palcic
keywords: affinity; barley; biochemical characterization; expression levels; kinetics; stability; starch synthases; substrate specificity