Starch Phosphorylation in Potato Tubers Proceeds Concurrently with de Novo Biosynthesis of Starch

Abstract

l h e in vivo phosphorylation of starch was studied in Solanum tuberosum cv Dianella and Posmo. Small starch granules contain 25% more ester-bound phosphate per glucose residue than large starch granules. l h e degree of phosphorylation was found to be almost constant during tuber development. lsolated tuber discs synthesize starch from externally supplied glucose at a significant rate. Tuber discs supplied with glucose and [3ZP]orthophosphate incorporate radiolabeled phosphorus into the starch. l h e level of 32P incorporation is proportional to the amount of starch synthesized. l h e incorporation of 32P from orthophosphate is correlated to de novo synthesis of starch, since the incorporation of 32P is diminished upon inhibition of starch synthesis by fluoride. Based on the amount of ['4C]glucose phosphate isolated after hydrolysis of purified starch from tuber discs incubated in the presence of [U-'4C]glucose, approximately 0.5% of the glucose residues of the de novo-synthesized starch are phosphorylated. This value is in general agreement with the observed levels of phosphorus in starch accumulated during tuber development. Thus, the enzyme system responsible for starch phosphorylation is fully active in the isolated tuber discs, and the starch phosphorylation proceeds as an integrated part of de novo starch synthesis.

Phosphorylation of Transitory Starch Is Increased during Degradation

“…Likewise, in potato leaves, starch phosphorylation occurs also during biosynthesis, as indicated by the similar levels of starch-bound Glc6P in light and darkness (Table I). Thus, transitory starch is phosphorylated during synthesis, as it has previously been documented for reserve starch in potato amyloplasts (Nielsen et al, 1994;Wischmann et al, 1999). However, here we show that during breakdown of transitory starch, phosphorylation occurs at an even higher rate.…”

“…However, experimental evidence is lacking. Nielsen et al (1994) have shown that potato tuber starch is phosphorylated during biosynthesis. This result, in combination with the more recent findings (Lorberth et al, 1998;Yu et al, 2001;Ritte et al, 2002), suggests that phosphate incorporation during starch synthesis somehow determines the degradability of the starch particle.…”

Glucan, water dikinase phosphorylates crystalline maltodextrins and thereby initiates solubilization

“…Under in vivo conditions, phosphorylation of native starch particles occurs both during net biosynthesis (Nielsen et al, 1994) and net degradation, but, at least in chloroplasts, the rate of phosphorylation is higher during net starch mobili-(a) (b) (c) Figure 5. MALDI MS analysis of purified phosphoglucans.…”

“…For enzymatic G-6-P quantification, the GWD assay mixtures were hydrolyzed in 0.7 N HCl (4 h at 95°C). Following neutralization with 0.7 N NaOH, G-6-P was quantified enzymatically (Nielsen et al, 1994). For TLC analyses, samples were hydrolyzed in 2 M TFA (2 h at 95°C), dried under vacuum, dissolved in water and dried again.…”

Simultaneous antisense inhibition of two starch-synthase isoforms in potato tubers leads to accumulation of grossly modified amylopectin

“…The amylose content of the purified starch was measured by two different methods ; first by quantitatively precipitating the amylopectin using the lectin Concanavalin A according to the instructions from a kit (Megazyme International), and secondly by an iodine-binding assay [18]. The glucose 6-phosphate content of the starch was measured using the enzyme glucose 6-phosphate dehydrogenase after acid hydrolysis of the starch [19].…”