109. Hayashi, T., Read, S. M., Bussell,
J., Thelen, M., Lin, F. C., Brown, Jr. R. M., and D. P. Delmer.
1987. UDP-Glucose (1-3)-b-Glucan
synthesis from mung bean and cotton. Plant Physiology 83:1054-1062.
109. Abstract
A re-examination of the kinetic properties
of UDP-glucose: (13--glucan (callose) syntheses from mung bean
seedlings (Vigna radiata) and cotton fibers (Gossypium
hirsutum) shows that these enzymes have a complex interaction
with UDP-glucose and various effectors. Stimulation of activity
by micromolar concentrations of Ca2+ and millimolar
concentrations of -glucosides or other polyols is highest at low
(<100 micromolar) UDP-glucose concentrations. These effectors
act both by raising the Vmax of the enzyme, and by
lowering the apparent Km for UDP-glucose from >1
millimolar to 0.2 to 0.3 millimolar Mg2+ markedly enhances
the affinity of the mung bean enzyme for Ca+2 but not
for -glucoside; with saturating Ca2+, Mg2+
only slightly stimulates further production of glucan. However,
the presence of Mg2+ during synthesis, or NaBH4
treatment after synthesis, changes the nature of the product from
dispersed, alkali-soluble fibrils to highly aggregated, alkali-insoluble
fibrils. Callose synthesized in vitro by the Ca2+,
-glucoside-activated cotton fiber enzyme, with or without Mg2+,
is very similar in size to callose isolated from cotton fibers,
but is a linear (13)--glucan lacking the small amount of branches
at C-O-6 found in vivo. We conclude that the high degree
of aggregation of the fibrils synthesized with Mg2+
in vitro is caused either by an alteration of the glucan
at the reducing end or, indirectly, by an effect of Mg2+
on the conformation of the enzyme. Rate-zonal centrifugation
of the solubilized mung bean callose synthase confirms that divalent
cations can affect the size or conformation of this enzyme.