2001 AACC Annual Meeting Charlotte, North Carolina October 14-18, 2001 Charlotte Convention Center

To establish a simple model system to study glycogen/starch biosynthesis, we investigated mechanism of glycogen biosynthesis in a cyanobacterium Synechocystis sp. PCC6803. Individual glycogen synthases (GSI and II) and branching enzyme (BE) were disrupted by homologous recombination conferring kanamycin resistance. Insertion of antibiotic resistance gene to disrupt each gene was confirmed by DNA amplification and Southern hybridization. Branch-structure of glycogen determined by high-performance anion-exchange chromatography (HPAEC) revealed that there was prominent increase in intermediate size chains of DP8-18 in GSI-deficient mutant compared to wild type. GSII-deficient mutant strain produced glycogen similar to wild type, with only a slightly higher proportion of short chains (DP4-11). Results indicated that the GS isoforms in Synechocystis sp. PCC6803 had different reaction specificities on glycogen biosynthesis. The analysis of glycogen content and structure of mutant strain showed that wild-type strain produced 34 mg glycogen/g wet cell mass and 74% of glycogen was in a water-soluble form. The BE-deficient mutant accumulated 16 mg glycogen/g wet cell mass and most of the glycogen (86%) was in a water-insoluble form. The insoluble glucan was retrograded linear molecules with negligible branches.