Fructan biosynthesis in excised leaves of Lolium temulentum L.

VI. Optimization and stability of enzymatic fructan synthesis

Authors

  • ANDREW J. CAIRNS,

    1. Environmental Biology Department, AFRC Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, SY23 3EB, UK
    Search for more papers by this author
  • JENNIFER E. ASHTON

    1. Environmental Biology Department, AFRC Institute of Grassland and Environmental Research, Plas Gogerddan, Aberystwyth, SY23 3EB, UK
    Search for more papers by this author

SUMMARY

A crude enzyme preparation from excised illuminated leaves of Lolium temulentum L. catalyzed the de novo synthesis of fructan of apparent degree of polymerization 3–20.

In the absence of sucrose, the fructan synthetic activity (FSA) was labile at temperatures above 10 °C, but could be stored at 5 °C for 6 h with the loss of only 20% of the initial activity. Sucrose stabilized the FSA such that high rates of fructan synthesis continued for at least 6 h at 30 °C. By comparison, in the absence of sucrose 80% of the FSA was lost in 2 h at 30 °C.

The FSA was maximal at pH 6·0 in citrate-phosphate buffer and at 43 °C. It was not possible to saturate the FSA even at 1500 mol m−3 sucrose, or to calculate meaningful kinetic parameters for enzymatic fructan synthesis. 150 mill m−3 sucrose was the highest substrate concentration practically feasible and imposed an arbitrary maximum concentration for use in enzyme assays. Pyridoxal-hydrochloride was found to reduce invertase activity by 40%, but also inhibited FSA by 20% at 1500 mol −3 sucrose and pH 6.0.

Ancillary

Advertisement