Molecular and biochemical characteristics of the inulosucrase HugO from Streptomyces viridochromogenes DSM40736 (Tü494)

Polyfructans are synthesized from sucrose by plants (mostly inulin) and by both Gram-negative and Gram-positive bacteria (mostly levan). In the phylum Actinobacteria only levan synthesis by Actinomyces species has been reported. We have identified a putative fructansucrase gene (hugO) in Streptomyces viridochromogenes DSM40736 (Tü494). HugO was heterologously expressed and biochemically characterized. HPSEC-MALLS and 2D-1H-13C nuclear magnetic resonance (NMR) spectroscopy analysis showed that the fructan polymer produced in vitro has an Molecular Weight of 2.5*107 Da and is an inulin that is mainly composed of (β2-1)-linked fructose units. This is the first report of a fructansucrase from Streptomyces and an inulosucrase from Actinobacteria. Database searches showed that fructansucrases clearly occur more widely in streptomycetes. Analysis of the active site of HugO and other actinobacterial Gram-positive fructansucrases revealed that their +1 substrate-binding sites are conserved, but are most similar to those in Gram-negative fructansucrases. HugO also resembles Gram-negative fructansucrases in not requiring calcium ions for activity. The origin and properties of HugO and other actinobacterial fructansucrases thus clearly differ from those of previously characterized Gram-positive fructansucrases.