Botrytis cinerea is known as a
β-(1-3)(1-6)-D-glucan-overproducer with an
exceptionally high degree of β-(1-6)-branches in the glucan. To
localize and characterize the glucan synthase, the incorporation of
[14C]glucose, activated by uridyl diphosphate
and incubated with mycelial extracts, into ethanol-precipitable
material was studied. Whereas crude extracts were found to produce
α- and β-glucans, a pure β-(1-3)-D-glucan
was formed by the membrane fraction. This was monitored by the
complete cleavage into glucose with a purified β-(1-3)-glucanase
and by the detection of laminaribiose, a β-(1-3)-linked dimer of
glucose, in incomplete digestions. Degradation products with
β-(1-6)-linkages, e.g. gentiobiose, which is found after
degradation of the native polymer, were not detectable.
β-(1-3)-glucan synthase activity was optimal at 22 °C and pH
7·2 with a Km of 0·8
mM and a Vmax of 0·24 mU
mg−1 protein. GTP
(Ka = 4·2 μM),
cellobiose,
BSA and EGTA enhanced the reaction
whereas UDP (Ki = 0·45 mM)
and
Ca2+ inhibited it.