| dc.description.abstract | Proline oxidation appears to constitute a major energy produring
pathway in flight and coxal muscle of various dung beetles
(Scarabaeidae: sub-family Coprine). This was concluded from the
fact that activities likely to be assocIated with a proline energy
metabolism such as proline dehydrogenase, alanine aminotransferase
and NAD-malic enzyme, were unexpectedly high in both flight and
coxal muscle of Coprine beetle, as was found for the better known,
proline utilising tsetsefly. In contrast, these enzyme activities
were relatively low in various other insects, all claimed to be
either lipid and/or carbohydrate utilisers. Major glycolytic
enzymes were relatively low in Coprine beetles and the tsetsefly,
in contrast to various other known carbohydrate utilising insects,
in which these activities were high. This suggests that carbohydrate
breakdown might not be important as an energy supplying
pathway in this beetle. It is noteworthy that NAD-malic enzyme
and alanine aminotransferase were mitochondrial in keeping with
a functi6nal role in proline metabolism.
Mitochondria from both flight and coxal muscle of Coprine
beetles oxidised proline faster than any other substrate tested.
This suggests that-proline constitutes a major physiological substrate.
In contrast, these mitochondria oxidised pyruvate and lipids
at negligible rates. Presumably, beetles are unable to utilise carbohydrates
and lipids directly. However; mitochondria from flight
muscle of "starved" beetles oxidised glycerophosphate faster
than proline and contained higher glycolytic enzyme activities.
It is possible therefore that the flight muscle tissue switches
to a carbohydrate based metabolism under "starved" conditions.
Flight and coxal muscle NAO-malic enzymes were purified
using the same procedure and their properties investigated under
identical conditions. Specific activities of 28 units/min/mg protein
and 25 units/min/mg protein were obtained for flight and
coxal muscle enzymes respectively. A similar molecular weight of
250 - 260 x 103 was estimated for the flight and coxal muscle
enzymes. The two enzymes had similar electrophoretic and kinetic
properties.
Catharsius NAD-malic enzyme had complete requirement for
Mn2+ or Mg2+ for activity and exhi bited co-operativity wi th respect
to both substrates. With malate as the variable substrate, the '.
x degree of sigmoidicity and the Krn values increased with pH. Succinate
activated the enzyme by increasing the affinity for malate
and tended to abolish co-operativity, as well as lowering the Km
values. Fumarate al~o activated the enzyme but unlike succinate,
high concentrations were strongly inhibitory.
The enzyme had relatively high Km values for both substrates,
probably as an adoptation to fit it for the control of mitochondrial
ma late utilisation. It was maximally 30 - 40% as active w ith
NAOP as the NAO-linked activity, and had similar affinity for the
coenzymes.
With succinate present, linear primary reciprocal initial
velocity-substrate plots were obtained. From the patterns of
these plots, it was deduced that the Cartharsius enzyme followed
a fixed order of substrate binding mechanism with malate binding
before NAD. Inhibition patterns 9btained with NADH and bicarbonate
were also in keeping with such a mechanism.
A Ca2+ dependent OAA decarboxyl ase acti vity was puri fied
together with NAD-malic enzyme activity and was probably attributable
to the same protein.
The reaction catalysed by NAD-malic enzyme was slightly
reversible apparently with high pyruvate and bicarbonate and its
unlikely that reductive carboxylation has any physiological significance
in Corpine beetles.
The properties of the Catharsius NAD-malic enzyme were
compared with those of otherNAD- and NADP-malic enzymes. The
relationship of velocity and malate concentration and the modification
by pH, succinate, ATP and metal activator were considered
withi n the concept of the monod Wyman Changeux (MWC) model | en |
