A fructan:fructan 1-fructosyltransferase (1-FFT) was purified from edible burdock for the first time, and a cDNA encoding 1-FFT was isolated from the plant. Two 1-FFTs named 1-FFTa and 1-FFTb were purified from extract of edible burdock by ammonium sulfate precipitation, followed by chromatographies on DEAE-Sepharose CL-6B, Toyopearl HW-55S and Sephadex G-100 columns. Inulin-type fructan such as nystose and fructooligosaccharides with higher DP were produced from 1-kestose by purified 1-FFTs. The general properties of both purified enzymes were very similar to each other. The purified 1-FFTa and 1-FFTb showed a single band by native PAGE and two bands, relative molecular masses of about 46,000 and 17,500 for 1-FFTa and 46,000 and 17,000 for 1-FFTb, by SDS-PAGE, respectively. The N-terminal sequences of the 46,000 peptides of both enzymes were the same, and those of the 17,500 and the 17,000 peptides were also identical. Based on the sequences, the 1-FFT cDNA named
alft1 was cloned. The
alft1 encoded a polypeptide of 617 amino acids. The relative molecular mass and p
I of the mature protein region of deduced polypeptide were calculated to be 60,213 and 4.89, respectively. The characteristics of recombinant protein produced by
Pichia pastoris closely resembled those of the native 1-FFTs purified from edible burdock. In this study, we demonstrated that
alft1 encoding edible burdock 1-FFT was involved in the elongation of fructosyl chains of fructooligosaccharides in edible burdock.
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