DISPROPORTIONATING ENZYME DEFINES A NOVEL STEP REQUIRED FOR NORMAL STARCH GRANULE BIOSYNTHESIS

Abstract

International audienceRecently, debranching enzymes were shown to be required during starch biosynthesis either for shaping amylopectin structure or for preventing synthesis of abnormal soluble polysaccharides in the plasmid stroma. We now describe a novel mutation in Clamydomonas which results in significantly reduced granular starch deposition displaying major modifications in structure. This defect simultaneously leads to the accumulation of linear malto-oligosaccharides. The mutation co-segregates with the absence of an -1,4 glucanotransferase , known as disproportionating enzyme (D-enzyme). We have cloned the gDNA and cDNA sequences corresponding to the Clamydomonas Reinhardtii -1,4 glucanotransferase.We demonstrate co-segregation between allele specific variations evidenced through Southern blots using the -1,4 glucanotransferase gDNA as a probe and the corresponding genotype at the STA11 locus. These results establish STA11 as the locus encoding D-enzyme.While the D-enzyme was known to transfer -1,4 linked chain segments between small-size acceptor and donor malto-oligosaccharides, we show that the missing enzyme readily catalyses the incorporation of oligosaccharides into amylopectin external chains in vitro.We therefore propose that D-enzyme plays a key role in starch biosynthesis by efficiently clearing those malto-oligosaccharides released by debranching enzyme. Oligosaccharide clearance can proceed by either reinserting spliced glucans into external chains of the putative maturing preamylopectin molecule or by facilitating their hydrolytic or phospholytic degradation