Granule‐bound starch synthase I

Abstract

Starch defines a semicrystalline polymer made of two different polysaccharide fractions. The A-and B-type crystalline lattices define the distinct structures reported in cereal and tuber starches, respectively. Amylopectin, the major fraction of starch, is thought to be chiefly responsible for this semicrystalline organization while amylose is generally considered as an amorphous polymer with little or no impact on the overall crystalline organization. STA2 represents a Chlamydomonas reinhardtii gene required for both amylose biosynthesis and the presence of significant granule-bound starch synthase I (GBSSI) activity. We show that this locus encodes a 69 kDa starch synthase and report the organization of the corresponding STA2 locus. This enzyme displays a specific activity an order of magnitude higher than those reported for most vascular plants. This property enables us to report a detailed characterization of amylose synthesis both in vivo and in vitro. We show that GBSSI is capable of synthesizing a significant number of crystalline structures within starch. Quantifications of amount and type of crystals synthesized under these conditions show that GBSSI induces the formation of B-type crystals either in close association with pre-existing amorphous amylopectin or by crystallization of entirely de novo synthesized material.Keywords: starch; amylose synthesis; granule-bound starch synthase; Chlamydomonas reinhardtii; in vitro synthesis.Starch accumulates in plants as a complex granular mixture of a-glucans (a-1,4-linked and a-1,6-branched) consisting chiefly of amylopectin and amylose. In amylopectin, the major fraction is composed of small-size a-1,4-linked chains that are clustered together by the presence of 5% a-1,6 linkages [1] (starch structure reviewed in [2] and [3]; starch metabolism reviewed in [4]). Amylose is composed of longer chains with less than 1% a-1,6 branches. Plant starch can be further distinguished from glycogen by the presence of highly ordered parallel arrays of double helical glucans (reviewed in [5]). The origin of these arrays resides in the close packing of the a-1,6 linkages at the root of the unit amylopectin cluster. The 9 nm size of each repetitive unit or cluster is conserved throughout the plant kingdom [6]. Two major types of crystalline organization have been documented so far in native starch granules. A-type powder diffraction patterns can be recovered from most cereal endosperm and Chlamydomonas reinhardtii starches while B-type structures were reported for tuber starches or high amylose starches from mutants of algae and cereals. It is generally assumed that amylopectin plays a major role in establishing the crystalline organization of starch. Indeed, amylosedefective mutants or antisense constructs of maize and potato accumulate normal amounts of starch with the same A-or B-type granule organization and similar crystallinities to the corresponding wild-type references. In addition, starches with elevated amylose content are generally less crystalline suggesting that ...

The barley amo1 locus is tightly linked to the starch synthase IIIa gene and negatively regulates expression of granule-bound starch synthetic genes

“…The effect of the sex 6 mutation was to decrease amylopectin synthesis predominantly (75% reduction on a per caryopsis basis) while amylose synthesis is decreased by 28% (Morell et al ., 2003). Given that amylose synthesis requires the granular matrix to be present (Wattebled et al ., 2002), the impact of the loss of SSIIa through the sex 6 mutation on amylose synthesis may be a secondary consequence of the major decrease in amylopectin synthesis in this mutant. It is evident that the high amylose content of the starch of s ex 6 mutants is a consequence of the differential inhibition of amylopectin synthesis.…”

Systems-Level Analysis of Nitrogen Starvation-Induced Modifications of Carbon Metabolism in a Chlamydomonas reinhardtii Starchless Mutant

“…reinhardtii, a unicellular chlorophyte alga containing a pyrenoid within its single chloroplast representing the site of starch accumulation and ribulose-1,5-bis-phosphate carboxylase/ oxygenase function, is an excellent reference system for addressing this issue (Harris, 2001). A draft genome sequence (http://www.phytozome.net/search.php for the version 5 assembly), resources for transcriptomics, proteomics, metabolomics (Bölling and Fiehn, 2005;May et al, 2008;Atteia et al, 2009;Rolland et al, 2009;Mühlhaus et al, 2011;Urzica et al, 2012;Wang et al, 2012), and well-characterized mutants in starch metabolism are available (Ball et al, 1991;Mouille et al, 1996;Wattebled et al, 2002Wattebled et al, , 2003. In addition, the growth medium and nutrient requirements are well defined, and there is considerable ongoing effort to discover and understand the operation of carbon metabolism pathways under different trophic situations, including anaerobiosis, respiratory, and phototrophic conditions (Kropat et al, 2011;Brueggeman et al, 2012;Catalanotti et al, 2012;Fang et al, 2012;Johnson and Alric, 2012).…”

Activity and expression of ADP-glucose pyrophosphorylase during rhizome formation in lotus (Nelumbo nucifera Gaertn.)

“…The second category of lotus cultivars is characterized by high starch content and low water content. The high content of granular starch gives the rhizome a soft texture, pliability, and increases the viscosity of root-derived products (Wattebled et al 2002). …”