Orthologs of Branching Enzymes from Cyanobacteria Accumulating Distinct Types of α-Glucans Share Common Reaction Product Specificity

Abstract

Cyanobacteria generally accumulate glycogen in their cells as a photosynthetic product. Interestingly, several unicellular diazotrophic species accumulate insoluble branched polysaccharide called cyanobacterial starch. Branching enzymes (BEs) belonging to glycoside hydrolase family 13 are universally found in the phylum cyanobacteria and are key enzymes in determining the branching pattern of polysaccharides. Many of the glycogen-producing cyanobacteria possess a single BE isozyme (BE1), while multiple BE isozymes (BE1, BE2, and BE3) are present in cyanobacterial starch-producing strains. A previous study suggested that the coexistence of three BE isozymes is essential for the trait of cyanobacterial starch-production. In this study, to obtain clues regarding the significance of the coexistence of the multiple isozymes, biochemical characterization using 11 purified recombinant BEs from both glycogen- and cyanobacterial starch-producing strains was performed. The BE1 and BE2 isozymes produced glucan chains with degree of polymerization (DP) 6 and 7 specifically, while BE3 isozymes produced short (DP 5-12) and long chains (DP 30-40) slightly. The BE1 and BE2 isozymes showed high activity, but those of BE3 isozymes were significantly low. The BE1 isozyme from cyanobacterial starch-producing Cyanobacterium sp. CLg1 showed markedly low activity. The BE1 and BE2 isozymes form cyanobacterial starch-producing Rippkaea orientalis PCC 8802 lacking BE3 isozyme shared similar reaction product specificity. These results suggested that the presence of the three isozymes is not essential and the roles of BE isozymes may vary depending on cyanobacterial species. These findings should deepen our understanding of the significance of BE isozymes in the biosynthesis of cyanobacterial starch.