The purpose of this work is to identify some of the mechanisms
regulating the conversion of assimilates to starch during endosperm
development in wheat.
Since environmental conditions affect the rate of grain
development, a standard system for ageing developing caryopses was
devised. The system was based on the morphological changes
accompanying caryopsis development, under field conditions, from
anthesis to harvest-ripeness. Accordingly, because the wheat
caryopsis passes through similar developmental changes which occur
at approximately the same relative time regardless of the time
scale, the tissues of the caryopsis, e.g. the endosperm, could be
standardised according to this system. This ensured that tissues at
the same stage of development could be compared, even when grown
under different environmental conditions, e.g. under glass.
Sucrose is the principal transported sugar in wheat and was found
to be the most abundant sugar in developing wheat endosperms.
Glucose and fructose were present in lower amounts and in different
relative quantities, with glucose declining throughout endosperm
development. The higher quantities of fructose were consistent with
low levels of endosperm invertase activity and much higher
activities of endosperm sucrose synthase.
Endosperm sucrose synthase activity reached an apparent maximum
catalytic rate during the period of rapid dry weight accumulation.
The levels of UDP in developing endosperms also reached a maximum
at this time. The curve of UDP-dependent sucrose synthase initial
reaction velocities was sigmoidal, thus endosperm UDP levels may
regulate the catabolism of sucrose.
Levels of ADP in developing wheat endosperms were higher than UDP
but UDP-glucose was present in amounts approximating to twice those
of ADP-glucose. This implies that ADP-dependent sucrose synthase
activity was not predominant in the catabolism of sucrose in
developing wheat endosperms.
Hexose sugars require to be in the form of hexose phosphates prior
to further metabolism by either glycolytic enzymes or ADP- and/or
UDP-glucose pyrophosphorylases. G6P was present in consistently
higher amounts than either G1P or F6P suggesting rapid
phosphorylation of glucose by hexokinase. Levels of G1P rose to a
maximum during endosperm dry weight accumulation and were quite
adequate to account for the measured rates of maximum velocity for
endosperm ADP-glucose pyrophosphorylase. G1P may have been formed
by the catabolism of UDP-glucose by the PPi-dependent UDP-glucose
pyrophosphorylase reaction. This enzyme activity was found to be
5-7 times higher in developing wheat endosperms than either sucrose
synthase or ADP-glucose pyrophosphorylase.
Differences were observed in the properties of ADP-glucose
pyrophosphorylases from endosperm and leaf tissues suggesting that
the control mechanisms differed between tissues. Both enzymes were
partially purified by ammonium sulphate fractionation and the
precipitates stored in 85 per cent ammonium sulphate. The endosperm
enzyme was stable in the unfractionated extract and in the stored
precipitate but, on subsequent dialysis of the stored precipitate,
rapidly lost activity, with a half-life of about 4h. The dialysed
activity was dependent on MgCl^ and was partially stabilised by Pi
but was not activated by 3-PGA. The leaf enzyme was stable to
fractionation by ammonium sulphate and to storage and dialysis but
both the crude tissue and partially purified activities were
dependent on the presence of 3-PGA.
PPi was found to be an efficient inhibitor of endosperm ADP-glucose
pyrophosphorylase while Pi was not, implying that PPi must be
removed from the site of ADP-glucose synthesis or hydrolysed. If
the generation of G1P for ADP-glucose synthesis is a result of
PPi-dependent catabolism of UDP-glucose then PPi/Pi metabolism may
regulate starch biosynthesis in developing wheat endosperms.