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

Abstract

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.

Highly porous core–shell chitosan beads with superb immobilization efficiency forLactobacillus reuteri121 inulosucrase and production of inulin-type fructooligosaccharides

“…Inulosucrase has been identified in only a few bacterial species such as Leuconostoc citreum , 13 Lactobacillus reuteri , 14 Lactobacillus johnsonii , 15 Lactobacillus gasseri , 16–18 and Streptomyces viridochromogenes . 19 Structural analysis shows that bacterial inulosucrases have low structural similarity when compared to fungal β-fructofuranosidase. In contrast, they are closely related to levansucrase (E.C.…”

High-yield production and purification of prebiotic inulin-type fructooligosaccharides

“…Crude InuGB-V3 exhibited rapid sucrose conversion at a broad pH spectrum (pH 3.5-6.0) while increased sucrose hydrolysis was observed at lower pH values. A pH optimum in the slightly acidic to neutral range was shown for many inulosucrases originating from lactic acid bacteria (Olivares-Illana et al 2002;Hijum et al 2002;Anwar et al 2008Anwar et al , 2010Ni et al 2018Ni et al , 2020, as well as for inulosucrases from Streptomyces viridochromogenes (Frasch et al 2017), Streptococcus mutans (Heyer et al 1998), and Bacillus macerans (Kim et al 1998). In addition, Ni et al (2018) showed for a variant of the inulosucrase from L. gasseri that the hydrolysis ratio increased at pH values below optimum.…”

Conserved Calcium-Binding Residues at the Ca-I Site Involved in Fructooligosaccharide Synthesis by Lactobacillus reuteri 121 Inulosucrase

“…Inulosucrase (E.C. 2.4.1.9) is a fructosyltransferase, which is found in bacterial species such as Leuconostoc citreum, Lactobacillus reuteri, Lactobacillus johnsonii, Lactobacillus gasseri, − and Streptomyces viridochromogenes . This enzyme converts sucrose to inulin-type fructooligosaccharide (IFOS) and β-2,1 fructan, a well-known prebiotic used in the food and pharmaceutical industries.…”