M. Runquist et Nj. Kruger, Control of gluconeogenesis by isocitrate lyase in endosperm of germinatingcastor bean seedlings, PLANT J, 19(4), 1999, pp. 423-431
The aim of this work was to quantify the contribution of isocitrate lyase t
o the control of gluconeogenesis in endosperm from 4-day-old castor bean se
edlings. The approach was based on metabolic control analysis following sel
ective inhibition of enzyme activity. Both 3-nitropropionate and itaconate
decreased the proportion of either [1-C-14]acetate or [2-C-14]acetate conve
rted to sucrose, and increased the proportion metabolized through the trica
rboxylic acid cycle. Kinetic analysis of isocitrate lyase activity from end
osperm revealed that itaconate is a pure uncompetitive inhibitor (K-i' = 11
.9 +/- 0.98 mu M) with respect to isocitrate. In contrast, 3-nitropropionat
e is a slow, tight-binding inhibitor. The half-time for inhibition of isoci
trate lyase by 3-nitropropionate was 5-10 min, whereas the half-time for re
activation was in excess of 10h. Incubating endosperm in 3-nitropropionate
resulted in a concentration-dependent decrease in isocitrate lyase activity
that remained stable in tissue extracts for at least 4h. From a comparison
of the extent of in situ inactivation of isocitrate lyase by 3-nitropropio
nate and the effect of this compound on the rate of sucrose production from
[2-14C]acetate, the flux control coefficient of isocitrate lyase on glucon
eogenesis from acetate in castor bean endosperm was calculated to be 0.66+/
-0.09. It is concluded that isocitrate lyase activity is quantitatively imp
ortant in the control of gluconeogenic flux, and suggested that development
al changes in the amount of this enzyme may be an important factor in deter
mining the conversion of lipid to sugar in young castor bean seedlings.