Identification of a novel DFA I-producing inulin fructotransferase from Streptomyces davawensis

In this work, a novel gene encoding DFA I-forming inulin fructotransferase (IFTase) from Streptomyces davawensis SK39.001 was cloned and expressed in Escherichia coli. The enzyme was purified, identified, and characterized. The results showed that this IFTase (DFA I-forming) is a trimer (molecular weight of 125 KDa) consisting of three identical subunits (the molecular weight as assayed by SDS-PAGE was approximately 40 KDa). At pH 5.5 and 40 °C, the maximum specific activity (approximately 100 U mg⿿1) was achieved. Moreover, the enzyme was stable up to 70 °C. Km and Vmax were 2.89 ± 0.2 mM and 1.94 ± 0.9 mM min⿿1, respectively. For exploring putative active sites and probable catalytic mechanisms, homology modelling and molecular docking methods after site-directed mutagenesis were applied to IFTase (DFA I-forming). The results revealed that D183 and E194 were potential catalytic residues of the purified enzyme.