Purification and properties of a heat stable inulin fructotransferase (DFA III-producing) from Arthrobacter pascens T13-2

An inulin fructotransferase (DFA III-producing) [EC 2.4.1.93] from Arthrobacter pascens T13-2 was purified and the properties of the enzyme were investigated. The enzyme was purified from a culture supernatant of the microorganism 18.5-fold with a yield of 13.1% by Super Q Toyopearl chromatographies and a butyl Toyopearl chromatography. It gave a single band on SDS-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular mass of the enzyme was estimated to be 44 000 by SDS-PAGE and 79 000 by gel filtration and was therefore considered to be a dimer. The N-terminal amino acid sequence was determined as Ala-Gln-Asp-Ala-Lys-Ala-Gly-Pro-Phe-Asn-Ser-Pro-Asn-Thr-Tyr-Asp-Val-Thr. The enzyme showed maximum activity at pH 5.5–6.0. The optimum temperature for the enzyme activity was 50 °C. The enzyme was stable up to 75 °C. The enzyme activity was inhibited strongly by Hg2+, and inhibited slightly by Fe3+, and Zn2+. An immobilized enzyme was prepared using Chitopearl BCW3510 as a carrier. The immobilized enzyme was able to use eight times without a significant loss of the enzyme activity.