0165 -
Fig. 1. Ripe ackee fruit, Blighia sapida. The arillus surrounding the seeds is cooked and
eaten, for example as 'ackee and salt fish; When unripe the ari/li contain dangerous
amounts of hypoglycin and hypoglycin B. During ripening these compounds are
translocated into the seeds.
6147186/$02.00
187
TIPS - M a y 1986
CH2=C.C.C.COOH are hypoglycaemic (Fig. 2). Of these, pent-4enoate has been extensively investigated because it was once
believed to be an analogue of
MCPA. However its mechanism of
action differs from that of hypoglycin (see Refs 6, 7).
The complete mitochondrial oxidation of long-chain fatty acids
requires the successive action of
palmitoyl-CoA, general acyl-CoA
and butyryl-CoA dehydrogenases
as the carbon chain is shortened by
successive loss of acetyl units.
MCPA-CoA inactivates the latter
two enzymes so that long-chain
acyl-CoA esters are only oxidized
as far as butyryl-CoA. ButyrylCoA accumulated is then converted to butyrate and CoASH by a
high Km acyl-CoA hydrolase. This
partial oxidation by inhibited liver
mitochondria is clearly shown
with conditions where the acetyl
groups formed by ~-oxidation are
quantitatively converted to acetoacetate 9 (Fig. 4). The maximum
possible rate of generation of
acetyl units from fatty acids is also
decreased by about half. This
hypoglycin (L-(methylenecyclopropyl)alanine)
hypoglycin B (%'-glutamylhypoglycin)
CHz~C
\*
CH.CH2.CH(NH2).COOH
CH2
~H.CH2.CH.COOH
I
NH.CO.CH2.CH2.CH(NH2).COOH
CHa
methylenecyclopropylpyruvate (MCPP)
CH2=C /
methylenecyclopropylacetate (MCPA)
Dicarboxylic aciduria
Hypoglycin-poisoned rats excrete large amounts of glutaric
acid 1. Glutaryl-CoA is an intermediate in the degradation of
tryptophan and lysine. Inhibition
of glutaryl-CoA dehydrogenase by
MCPA-CoA, followed by deacylation of accumulated glutaryl-CoA
yields glutarate. Rats also excrete
CH2
CH2=C
CH.CH
*
2. C O . C O 2
CHz
CH2=C/
~ ~H.CH..CO~-
H2-~H.CO23-methylenecyclobutanecarboxylate
CHz=CH--CH 2
pent-4-enoate
CH2=CH.CH~.CH2.CO~-
Fig. 2. Hypoglycin and some related compounds. Hypoglycin is an L-amino acid, C~is asymmetric, and the structure is thought to be (+)(2S: 4S)-2-amino-4,5-methanohexo5-enoicacid. MCPP and MCPA (as its CoA-estor) are derived from hypoglycin in vivo (see Fig. 3). Most
preparations of hypoglycin contain up to 20% of leucine, pure hypoglycin is obtained by hydrolysis of hypoglycin B first separated from
leucine by ion-exchange chromatography. 3-Methylenecyciobutanecarboxylate is a synthetic analogue of hypoglycin. All these compounds
contain the grouping CH2 = CH C.C.COOH. MCPA-CoA (and possibly 3-methylenecyciobutanecarbonyI-CoA) appears to undergo
2-deprotonization by some acyI-CoA dehydrogenases to form suicide inhibitors. Pent-4-enoyI-CoA formed in vivo is a substrato for butyrylCoA dehydrogenase without inhibiting it, and is partly metabolized to 3-oxopent-4-enoyI-CoA which inhibits mitochondrial ~-oxidation by
inactivating the 3-oxoacyI-CoA thiolases.
188
hypoglycin _
2-oxoglutarate
glutamate ~'-
-- MCPP
NAD
~NADH+H+/
MCPA-CoA
MCPA
AMP + PPi ~
"
\ CoASH + ATP
-glycine
CoASH
MCPA-glycine
Fig. 3. Metabolism of hypoglycin. MCPA-CoA is formed in mitochond/~a by the enzymes which catalyse the degradation of branched-chain
amino acids. MCPA-CoA maybe hydrolysed to free MCPA which is partly reconverted to MCPA-CoA, or partly conjugated with glycine. The
enzymes catalysing these reactions are: I. Leucine-2-oxoglutarate aminotransferase in the cell cytosol 2. Branched-chain 2-oxoacid
dehydrogenase in the mitochondrial matrix 3. Glycine N-acylase in the matrix 4. Medium-chain acyI-CoA hydrolase in the matrix 5. ButyrylCoA synthase in the mattYx.
L
I
glycemld.ehyi.3"phosphate
NAD +
1,3-diphosphoglycerate
phosph~nop~mvate
cytoso]
tase 13.
J3-oxidation J
rm'toch ondn'a/raatr/x
I n h i b i t i o n of branched-chain fatty
acid o x i d a t i o n
The
branched-chain
amino
acids leucine, isoleucine and valine are transaminated to the corr e s p o n d i n g 2-oxoacids which are
then oxidatively decarboxylated to
isovaleryl-CoA, 2-methylbutyryl-
189
TIPS - M a y 1986
< palmitoyl-carnitine
(79)
430
~P~
;palmitoyl-carniline
oxygen uptake
T
400 n g - a t o m of 0 / ~
1
2min
Fig. 4. Experiment illustrating partial inhibition of mitochondrial ~-oxidatlon (from Ref. 9). A
rat liver mitochondrial fraction, 10 i~Mpaimitoyl-carnitine and 1.0 mM- MCPA were added
where indicated to 3.0 ml of medium at 30C containing 100 mM KCI, 2.5 mM phosphate,
5 mM MgCI2, 1.0 mM EDTA, 2.0 mM ADP (to maintain the maximum rate of electron
transport), tO mM malonata (to prevent oxidation of acetyl groups by the citrate cycle),
9 mg of defatted bowne serum albumin and 20 mM 3 (morpholino) propanesulphonate
(buffer), pH 7.2. Oxygen uptake was recorded potarographically. Rates of oxygen uptake
(ngatom O/mg of protein) are given in parenthesas, and the amount of oxygen consumed
by vertical arrows. PalmitoyI-CoA is germratdd in the mitochondrial matrix from added
palmitoyl-carnitine by the action of the camitine palmitoyltransferasas in the inner
membrane. In control incubations palmitoyI-CoA is quantitatively oxidized to acetoacetate
PalmitoyI-CoA + 702 - * 4Acetoacetate + CoASH
and endogenous respiration is largely suppressed during paimitoyl-camitine oxidation.
After incubation for 3 min with MCPA to allow formation of MCPA-CoA in the math'x, both
the rate and extent of oxidation of palmitoyI-CoA are decreased:
PalmitoyI-CoA + 602--~ 3Acetoacetate + Butyrate + CoASH
(see text). Similar results were obtained with even-chain acyl-carnitine asters (Ca to C14).
If MCPA was incubated with uncoupled mitochondria (in the presence of 0.2 pM
trifluormelhoxycarbonylcyanidephenylhydrazone, lOmM arsenate and 0.6mg of
valinomycin and absence of ADP) no inhibition was found of acyl-carnitine oxidation
indicating that prior A TP-dependent conversion of MCPA to MCPA-CoA is necessary.
Similar inhibitions are found in mitochondria prepared from rats which have been injected
with hypoglycin ~~.
190
TIPS - M a y 1986
[ 2 Rate of ]
3 Rateof
breakdown
of hepatic
andrenal
glycogen
4 Rateof
utilization
of glucose
5 Rateof
glycogen ]
- synthesis in] liverand I
other
[
tissues ]
6 Rateof
[ excretion
ofglucose
I
b y h y p o t h e r m i a in small animals
(see Ref. 6). A n attempt was made
.to get quantitative information
about glucose metabolism b y a
kinetic study of the rate of decline
of the specific radioactivity of
blood glucose following a bolus
i.v. dose of [U-14C,2-3H]glucose
16 h after administration of hypoglycin (100 mg kg -1 body-wt) to
fasted rats (body temperatures
were artificially maintained) 17. Results indicated that gluconeogenesis was strongly inhibited
since recycling of glucose through
the Cori cycle (resynthesis of glucose in the liver from lactate formed
from glucose in peripheral tissues
b y glycolysis) was virtually abolished and the w h o l e - b o d y content
of free glucose was decreased b y
70%. The rate of utilization of
glucose was decreased b y 70%,
rather than increased as expected if
fatty acid oxidation is impaired. It
was concluded that hypoglycaemia
resulted from an even greater
191
TIPS - M a y 1986
[]
[]
[]