title: Glucan affinity of starch synthase IIa determines binding of starch synthase I and starch-branching enzyme IIb to starch granules.
abstract: The sugary-2 mutation in maize (Zea mays L.) is a result of the loss of catalytic activity of the endosperm-specific SS (starch synthase) IIa isoform causing major alterations to amylopectin architecture. The present study reports a biochemical and molecular analysis of an allelic variant of the sugary-2 mutation expressing a catalytically inactive form of SSIIa and sheds new light on its central role in protein-protein interactions and determination of the starch granule proteome. The mutant SSIIa revealed two amino acid substitutions, one being a highly conserved residue (Gly522→Arg) responsible for the loss of catalytic activity and the inability of the mutant SSIIa to bind to starch. Analysis of protein-protein interactions in sugary-2 amyloplasts revealed the same trimeric assembly of soluble SSI, SSIIa and SBE (starch-branching enzyme) IIb found in wild-type amyloplasts, but with greatly reduced activities of SSI and SBEIIb. Chemical cross-linking studies demonstrated that SSIIa is at the core of the complex, interacting with SSI and SBEIIb, which do not interact directly with each other. The sugary-2 mutant starch granules were devoid of amylopectin-synthesizing enzymes, despite the fact that the respective affinities of SSI and SBEIIb from sugary-2 for amylopectin were the same as observed in wild-type. The data support a model whereby granule-bound proteins involved in amylopectin synthesis are partitioned into the starch granule as a result of their association within protein complexes, and that SSIIa plays a crucial role in trafficking SSI and SBEIIb into the granule matrix.
authors.affiliation: Department of Molecular and Cellular Biology, College of Biological Science, University of Guelph, Guelph, ON, Canada, N1G 2W1.
authors.forename: Fushan
authors.initials: F
authors.lastname: Liu
authors.forename: Nadya
authors.initials: N
authors.lastname: Romanova
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Lee
authors.forename: Regina
authors.initials: R
authors.lastname: Ahmed
authors.forename: Martin
authors.initials: M
authors.lastname: Evans
authors.forename: Elliot P
authors.initials: EP
authors.lastname: Gilbert
authors.forename: Matthew K
authors.initials: MK
authors.lastname: Morell
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Emes
authors.forename: Ian J
authors.initials: IJ
authors.lastname: Tetlow
mesh_terms: D015061:1,4-alpha-Glucan Branching Enzyme / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000483:Alleles; D000595:Amino Acid Sequence; D000687:Amylopectin / Q000737:chemistry; D005936:Glucans / Q000737:chemistry* / Q000235:genetics; D006006:Glycogen Synthase / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D013213:Starch / Q000737:chemistry* / Q000235:genetics; D013214:Starch Synthase / Q000737:chemistry* / Q000235:genetics; D003313:Zea mays / Q000201:enzymology
chemical_list: D005936:Glucans; D010940:Plant Proteins; D013213:Starch; D000687:Amylopectin; C111377:starch synthase II; D006006:Glycogen Synthase; D015061:1,4-alpha-Glucan Branching Enzyme; C116400:starch-branching enzyme IIb; D013214:Starch Synthase