Tracking sulfur and phosphorus within single starch granules using synchrotron X-ray microfluorescence mapping

Abstract : Background : Native starch accumulates as granules containing two glucose polymers: amylose and amylopectin. Phosphate (0.2-0.5%) and proteins (0.1-0.7%) are also present in some starches. Phosphate groups play a major role in starch metabolism while granule-bound starch synthase 1 (GBSSI) which represents up to 95% of the proteins bound to the granule is responsible for amylose biosynthesis. Methods : Synchrotron micro-X-ray fluorescence (mu XRF) was used for the first time for high-resolution mapping of GBSS1 and phosphate groups based on the XRF signal of sulfur (S) and phosphorus (P), respectively. Wild-type starches were studied as well as their related mutants lacking GBSS1 or starch-phosphorylating enzyme. Results : Wild-type potato and maize starch exhibited high level of phosphorylation and high content of sulfur respectively when compared to mutant potato starch lacking glucan water dikinase (GWD) and mutant maize starch lacking GBSSI. Phosphate groups are mostly present at the periphery of wild-type potato starch granules, and spread all over the granule in the amylose-free mutant P and S XRF were also measured within single small starch granules from Arabidopsis or Chlamydomonas not exceeding 3-5 mu m in diameter. Conclusions : Imaging GBSSI (by S mapping) in potato starch sections showed that the antisense technique suppresses the expression of GBSS1 during biosynthesis. P mapping confirmed that amylose is mostly present in the center of the granule, which had been suggested before. General significance: IMF is a potentially powerful technique to analyze the minor constituents of starch and understand starch structure/properties or biosynthesis by the use of selected genetic backgrounds.