Lehrstuhl für Biotechnologie, RWTH Aachen, Sammelbau Biologie, Worringer Weg 1, D-52074 Aachen, Germany.
Seasonal variation of fructan-β-fructosidase (FEH) activity and characterization of a β-(2-1)-linkage specific FEH from tubers of Jerusalem artichoke (Helianthus tuberosus)
Article first published online: 28 JUN 2008
Volume 135, Issue 2, pages 267–277, February 1997
How to Cite
MARX, S. P., NÖSBERGER, J. and FREHNER, M. (1997), Seasonal variation of fructan-β-fructosidase (FEH) activity and characterization of a β-(2-1)-linkage specific FEH from tubers of Jerusalem artichoke (Helianthus tuberosus). New Phytologist, 135: 267–277. doi: 10.1046/j.1469-8137.1997.00641.x
- Issue published online: 28 JUN 2008
- Article first published online: 28 JUN 2008
- (Received 1 April 1996; accepted 27 September 1996)
- Fructan-β-fructosidase (1-FEH; EC 18.104.22.168);
- fructan hydrolysis;
- Helianthus tuberosus (Jerusalem artichoke);
The fructan-β-fructosidase activity (1-FEH; EC 22.214.171.124) that degrades inulin in tubers of Helianthus tuberosus L. appears to be developmentally regulated; it was low in growing tubers but increased during dormancy and sprouting. In spite of relatively high 1-FEH activity in vitro, fructose concentration was very low in developing and dormant tubers and increased markedly only during sprouting. A fructan-β-fructosidase from such sprouting tubers was purified 41-fold to a single protein band on one-dimensional sodium dodecylsulphate-polyacrylamide gels. The purification procedure included ammonium sulphate precipitation, lectin-affinity chromatography on concanavalin A, anion-exchange and cation-exchange chromatography. The enzyme had an apparent molecular mass of 75000 measured by size-exclusion chromatography, and 79000 measured by one-dimensional sodium dodecylsulphate-polyacrylamide gel electrophoresis. It exhibited a high substrate specificity, hydrolysing terminal β-(2-1)-fructosyl-fructose-linkages in linear and branched fructan oligomers; β-(2-6)-linkages were hardly hydrolysed. Hydrolysis of inulin oligomers followed normal saturation kinetics: Km values for 1,1-kestotetraose and 1,1,1-kestopentaose were 8-3 mM and 12 mM, respectively. Fructosyl residues were hydrolysed from inulin oligomers by a multi-chain mechanism. The fructan-β-fructosidase showed optimal enzyme activity at pH 5–2, and it retained its full activity after pre-incubation for 1 h at up to 40°C. The release of fructose from 5 mm 1,1-kestotetraose was reduced by 25 % when 1-FEH was assayed in the presence of 10 mM sucrose. It is proposed that the inhibition of 1-FEH activity by sucrose is a mechanism for controlling fructan degradation in planta.
- Con A
degree of polymerization
FEH that hydrolyse predominantly β-(2-1)-linkages
fructan:fructan fructosyl transferase
high-performance anion-exchange chromatography with pulsed amperometric detection
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