title: Protein-protein interactions among enzymes of starch biosynthesis in high-amylose barley genotypes reveal differential roles of heteromeric enzyme complexes in the synthesis of A and B granules.
abstract: The present study investigated the role of protein phosphorylation, and protein complex formation between key enzymes of amylopectin synthesis, in barley genotypes exhibiting "high amylose" phenotypes. Starch branching enzyme (SBE) down-regulated lines (ΔSBEIIa and ΔSBEIIb), starch synthase (SS)IIa (ssiia(-), sex6) and SSIII (ssiii(-), amo1) mutants were compared to a reference genotype, OAC Baxter. Down-regulation of either SBEIIa or IIb caused pleiotropic effects on SSI and starch phosphorylase (SP) and resulted in formation of novel protein complexes in which the missing SBEII isoform was substituted by SBEI and SP. In the ΔSBEIIb down-regulated line, soluble SP activity was undetectable. Nonetheless, SP was incorporated into a heteromeric protein complex with SBEI and SBEIIa and was readily detected in starch granules. In amo1, unlike other mutants, the data suggest that both SBEIIa and SBEIIb are in a protein complex with SSI and SSIIa. In the sex6 mutant no protein complexes involving SBEIIa or SBEIIb were detected in amyloplasts. Studies with Pro-Q Diamond revealed that GBSS, SSI, SSIIa, SBEIIb and SP are phosphorylated in their granule bound state. Alteration in the granule proteome in ΔSBEIIa and ΔSBEIIb lines, suggests that different protein complexes are involved in the synthesis of A and B granules.
authors.affiliation: Department of Molecular and Cellular Biology, College of Biological Science, University of Guelph, Guelph, Ontario, N1G 2W1, Canada. Electronic address: zahmed@uoguelph.ca.
authors.forename: Zaheer
authors.initials: Z
authors.lastname: Ahmed
authors.affiliation: Department of Molecular and Cellular Biology, College of Biological Science, University of Guelph, Guelph, Ontario, N1G 2W1, Canada. Electronic address: itetlow@uoguelph.ca.
authors.forename: Ian J
authors.initials: IJ
authors.lastname: Tetlow
authors.affiliation: Food Futures National Research Flagship and Division of Plant Industry, CSIRO, Canberra ACT 2601, Australia. Electronic address: regina.regina@csiro.au.
authors.forename: Regina
authors.initials: R
authors.lastname: Ahmed
authors.affiliation: International Rice Research Institute, Manilla, Phillipines. Electronic address: m.morell@irri.org.
authors.forename: Matthew K
authors.initials: MK
authors.lastname: Morell
authors.affiliation: Department of Molecular and Cellular Biology, College of Biological Science, University of Guelph, Guelph, Ontario, N1G 2W1, Canada. Electronic address: memes@uoguelph.ca.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Emes
mesh_terms: D015536:Down-Regulation; D001467:Hordeum / Q000201:enzymology / Q000235:genetics*; D008854:Microscopy, Electron; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D020543:Proteome; D013213:Starch / Q000096:biosynthesis* / Q000648:ultrastructure
chemical_list: D009097:Multienzyme Complexes; D010940:Plant Proteins; D020543:Proteome; D013213:Starch
keywords: A and B granules; Barley; Enzyme complexes; Starch biosynthesis