Red algal starch metabolism : effects of endosymbionts and nutrient status on starch metabolising enzymes

Sammanfattning: Many different organisms occur as endosymbionts of red algae and while some are highly pathogenic others have little effect on the host. An endosymbiotic fungus, found in our culture of Gracilariopsis sp., was characterised and identified as a Chytridiomycete. It was named Thalassochytrium gracilariopsidis and probably belongs to a new order. Another group of endosymbionts is the parasitic red algae. Based on morphological characters it is suggested that they have evolved from their respective red algal host. ITS and 18S sequence data of a number of parasitic red algae established that the majority are more closely related to their host than to other closely related species.Infection with either the parasitic red alga Gracilariophila oryzoides or the endosymbiotic fungus increased the activity of the starch degrading enzyme (-1,4-glucan lyase in Gracilariopsis sp. The enzyme was examined regarding its kinetic characteristics and active-site amino acids. Similarly to several (-glucosidases, a carboxyl group was found to be important for the catalytic activity.Based on the unique composition and cytosolic location of floridean starch in red algae the starch synthesising enzymes in Gracilaria tenuistipitata were examined. From this alga a novel UDPglucose: starch synthase was isolated and characterised. The isolated enzyme is most active with UDPglucose as substrate and appears to be a homotetramer. Thus, it is biochemically more similar to glycogen synthases from animals and fungi than to starch synthases from plants. Regardless of the nutrient status of G. tenuistipitata, the activity with ADPglucose was at the most 3% of that with UDPglucose. Thus, UDPglucose appears to be more important as substrate for starch synthesis in G. tenuistipitata than is ADPglucose. From our data I suggest that the pathway of starch synthase in red algae has a eukaryotic rather than prokaryotic ancestry.During the purification of the starch synthase an HPLC assay for the quantification of starch synthase activity was developed that was less labour-intensive than classical activity determinations and which allowed the use of non-radioactive substrates.