Purification and Characterization of Inulin Fructotransferase (Depolymerizing) from Arthrobacter sp. A-6

Inulin fructotransferase (depolymerizing) (EC 2.4.1.93) was purified 34-fold from the culture broth of Arthrobacter sp. A-6 by using a combination of ammonium sulfate fractionation, DEAE-Sepharose CL-6B chromatography and Sephacryl S-200 gel filtration. The purified enzyme converts inulin into di-D-fructofuranose dianhydride III(DFA III) and small quantities of fructo-oligosaccharides. The temperature and pH optima of the enzyme were <TEX>$70^{circ}C$</TEX> and 6.0, respectively. Molecular weight of the enzyme was determined to be 49 kDa by 12<TEX>$%$</TEX> SDS-polyacrylamide gel electrophoresis, and 145 kDa by Sephacryl S-200gel filtration. This indicates that the functional inulin fructotransferase of Arthrobacter sp. A-6 has a homomeric trimer structure. The enzyme had an isoelectric point of pH 4.6. The N-terminal amino acid sequence of the purified enzyme subunit was Ala-Asp-Asn-Pro-Asp-Gly(ulcorner)-Ser-Asn-Met(or Glu)-Tyr-Asp-Val.