Significant increases in potato Starch-Synthase and Starch-Branching-Enzyme activities by dilution with buffer containing dithiothreitol and polyvinyl alcohol 50K

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.

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Phosphorylated α(1→4)Glucans as Substrate for Potato Starch-Branching Enzyme I

PLANT PHYSIOLOGY ◽

10.1104/pp.117.3.869 ◽

1998 ◽

Vol 117 (3) ◽

pp. 869-875 ◽

Cited By ~ 16

Author(s):

Anders Viksø-Nielsen ◽

Andreas Blennow ◽

Tom Hamborg Nielsen ◽

Birger Lindberg Møller

Keyword(s):

Potato Starch ◽

Branching Enzyme ◽

Starch Branching Enzyme ◽

Enzyme I

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Inactivation of rice starch branching enzyme IIb triggers broad and unexpected changes in metabolism by transcriptional reprogramming

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2014860117 ◽

2020 ◽

Vol 117 (42) ◽

pp. 26503-26512

Author(s):

Can Baysal ◽

Wenshu He ◽

Margit Drapal ◽

Gemma Villorbina ◽

Vicente Medina ◽

...

Keyword(s):

Amylose Content ◽

Starch Biosynthesis ◽

Starch Synthase ◽

Rice Starch ◽

Biosynthesis Pathway ◽

Branching Enzyme ◽

Starch Branching Enzyme ◽

Rice Endosperm ◽

Primary And Secondary Metabolites ◽

Genes Encoding

Starch properties can be modified by mutating genes responsible for the synthesis of amylose and amylopectin in the endosperm. However, little is known about the effects of such targeted modifications on the overall starch biosynthesis pathway and broader metabolism. Here we investigated the effects of mutating theOsSBEIIbgene encoding starch branching enzyme IIb, which is required for amylopectin synthesis in the endosperm. As anticipated, homozygous mutant plants, in which OsSBEIIb was completely inactivated by abolishing the catalytic center and C-terminal regulatory domain, produced opaque seeds with depleted starch reserves. Amylose content in the mutant increased from 19.6 to 27.4% and resistant starch (RS) content increased from 0.2 to 17.2%. Many genes encoding isoforms of AGPase, soluble starch synthase, and other starch branching enzymes were up-regulated, either in their native tissues or in an ectopic manner, whereas genes encoding granule-bound starch synthase, debranching enzymes, pullulanase, and starch phosphorylases were largely down-regulated. There was a general increase in the accumulation of sugars, fatty acids, amino acids, and phytosterols in the mutant endosperm, suggesting that intermediates in the starch biosynthesis pathway increased flux through spillover pathways causing a profound impact on the accumulation of multiple primary and secondary metabolites. Our results provide insights into the broader implications of perturbing starch metabolism in rice endosperm and its impact on the whole plant, which will make it easier to predict the effect of metabolic engineering in cereals for nutritional improvement or the production of valuable metabolites.

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Functional Analysis of Starch Metabolism in Plants

Plants ◽

10.3390/plants9091152 ◽

2020 ◽

Vol 9 (9) ◽

pp. 1152 ◽

Cited By ~ 1

Author(s):

Yong-Gu Cho ◽

Kwon-Kyoo Kang

Keyword(s):

Transcription Factors ◽

Starch Biosynthesis ◽

Starch Synthase ◽

Starch Metabolism ◽

Branching Enzyme ◽

Structural Genes ◽

Starch Branching Enzyme ◽

Fixed Carbon ◽

Debranching Enzyme ◽

Storage Organs

In plants, starch is synthesized in leaves during the day-time from fixed carbon through photosynthesis and is mobilized at night to support continued respiration, sucrose export, and growth in the dark. The main crops where starch is biosynthesized and stored are corn, rice, wheat, and potatoes, and they are mainly used as food resources for humankind. There are many genes that are involved in starch biosynthesis from cytosol to storage organs in plants. ADP-glucose, UDP- glucose, and glucose-6-phosphate are synthesized catalyzed by UDP-invertase, AGPase, hexokinase, and P- hexose-isomerase in cytosol. Starch composed of amylopectin and amylose is synthesized by starch synthase, granule bound starch synthase, starch-branching enzyme, debranching enzyme, and pullulanase, which is primarily responsible for starch production in storage organs. Recently, it has been uncovered that structural genes are controlled by proteins derived from other genes such as transcription factors. To obtain more precise information on starch metabolism, the functions of genes and transcription factors need to be studied to understand their roles and functions in starch biosynthesis in plants. However, the roles of genes related to starch biosynthesis are not yet clearly understood. The papers of this special issue contain reviews and research articles on these topics and will be a useful resource for researchers involved in the quality improvement of starch storage crops.

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