Download Beta-oxidation of docosahexaenoic acid and tetracosahexaenoic

Biochemical Society Transactions (200 I ) Volume 29, Part I
6 Cloning and functional expression of human peroxisomal
2,4-dienoyl-CoA reductase
K. De Nys, E. Meyhi, G. Mannaerts, P. Van Veldhoven
Katholieke Universiteit Leuven, Afdeling Farmacologie,
Herestraat 49, B-3000 Leuven
2,4 Dienoyl-CoA reductase (DCR) is an auxiliary beta-oxidation
enzyme required for the degradation of unsaturated fatty acids
with even-numbered double bounds. The enzyme activity is
present in both mitochondria and peroxisomes, but catalysed by
different proteins. The peroxisomal rat liver DCR, which was
cloned before by means of phage-display technology (Fransen et
al, 1999, BJ 340,561), was used as a query to recover sequences
coding for the human DCR from the human EST1 and genomic
DNA databases. A fragment of 900 bases from a human liver
cDNA library was amplified and subcloned into the pBadHis
vector.
The bacterially expressed His-tagged 2,4-dienoyl-CoA reductase
showed a clear activity not only towards 2-trans, 4-trans-hexadienoyl-CoA (sorboyl-CoA) and 2-trans,
4-trans-decadienoyl-CoA, but also towards 2-trans, +cis, 7-cis,
10-cis, 13-cis, 16-cis, 19-cis-docosaheptaenoyl-CoA.The latter
CoA-ester is produced during the beta-oxidation of docosahexaenoic acid. DCR activity towards docosaheptaenoyl-CoA
was inhibited in the presence of albumin whereas sorboyl-CoA
reduction was not influenced by albumin.
Hence, DCR is likely to be important for the peroxisomal degradation of polyunsaturated fatty acids.
A23
8 Beta-oxidation of docosahexaenoic acid and tetracosahexaenoic acid in isolated rat hepatocytes and human
fibroblasts
K. De Nvs, E. Meyhi, S. Asselberghs, G. Mannaerts,
P. Van Veldhoven
Katholieke Universiteit Leuven, Farmacologie, Herestraat 49,
3000 Leuven
The pathways involved in the synthesis and degradation of very
long chain polyunsaturated fatty acids are still unclear.
Peroxisomes have been implicated in the formation of docosahexaenoic acid, which would be formed by subjecting
tetracosahexaenoic acid, generated in the endoplasmic reticulum,
to one beta-oxidation cycle. In order to study this process, isolated human fibroblasts and rat hepatocytes were incubated with
[I -'4C]docosahexaenoic acid and [2-14C]tetracosahexaenoicacid.
Both docosahexaenoic acid and tetracosahexaenoic acid were
beta-oxidized by control human fibroblasts and isolated rat hepatocytes. In fibroblasts from a Zellweger patient, docosahexaenoic
acid and tetracosahexaenoic acid degradation could not be
detected. After clofibrate treatment, a 3-fold increase in oxidation
rates in rat hepatocytes was noted. Preincubation of hepatocytes
with tetradecylglycydic acid resulted in a significant inhibition of
the oxidation of docosahexaenoic acid, but not of tetracosahexaenoic acid.
Our results suggest that tetracosahexaenoic acid can be degraded
to docosahexaenoic acid. In human fibroblasts both fatty acids are
beta-oxidized, mainly in peroxisomes. In rat liver, tetracosahexaenoic acid seems to be a selective peroxisomal substrate while
docosahexaenoic acid is partly oxidized in mitochondria. The peroxisomal enzymes implicated in the breakdown of these fatty
acids are induced by hypolipidemic drugs.
7 Is a microsomal aldehyde dehydrogenase involved in the
degradation of 3-methyl branched fatty acids?
V. F o h , L. Van Vaeck, S. Asselberghs, G. Mannaerts,
P. Van Veldhoven, M. Casteels
Katholieke Universiteit Leuven, Farmacologie, Herestraat 49,
B-3000 Leuven
The primary products of the degradation of 3-methyl branched
fatty acids such as phytanic acid are formyl-CoA and a 2-methyl
branched fatty aldehyde. Since this alpha-oxidation process and
the subsequent beta-oxidation of the 2-methyl branched fatty acid
were clearly shown to be peroxisomal, the role of a microsomal
fatty aldehyde dehydrogenase (FALDH) in the conversion of 2methyl fatty aldehydes appears somewhat intriguing.
In order to investigate a putative peroxisome/ER transfer of
alpha-oxidation intermediates, degradation of 3-methyl-[2I4C]heptadecanoic acid was studied in intact cells. The release of
labeled ASM produced in consecutive alpha- and beta-oxidation
by cultured skin fibroblasts of patients with Sjogren-LarssonSyndrome (SLS), known to be deficient in fatty aldehyde
dehydrogenation, was 30 to 60 % of the amount released by
control fibroblasts. However, conversion of 2-methylpentadecanal into 2-methylpentadecanoic acid in homogenates of SLS
fibroblasts, as measured by GC, was only marginally impaired.
Measurement of degradation of 3-methyl-[2-'4C]heptadecanoic
acid in homogenates is difficult due to the need for different
incubation conditions for alpha- and beta-oxidation.
Hence, measuring consecutive alpha- and beta-oxidation could
not prove the exclusive involvement of a microsomal FALDH in
the degradation of 3-methyl branched fatty acids.
9 Arachidonate and docosahexaenoate could be biochemical
markers to differentiate between Crohn's disease and
ulcerative colitis.
D.C. Bitsanis', R. Heuschkelz, K. Ghebremeskel',
J. Walker-Smithz, and M.A. Crawford'.
'Institute of Brain Chemistry and Human Nutrition, University
of North London, 146-222Holloway Road, London N7 8DB,
UK, 2University Department of Paediatric Gastroenterology,
Royal Free Campus, London, NW3 2Pe UK.
We have investigated red cell membrane phospholipid fatty acid
composition in Crohn's disease (CD, n=lo)and ulcerative colitis
(UC, n=7) patients. The U C patients had lower levels of dihomo-1inolenic (203n-6, p<O.O1), arachidonic (AA, p=0.001), adrenic
(22:4n-6, p=0.002), docosapentaenoic (22:5n-3, p<O.O1) and
docosahexaenoic (DHA, pc0.05) acids in the erythrocyte
ethanolamine phosphoglycerides (EPG) compared to the C D
patients. Similarly, in the choline phosphoglycerides (CPG), the
levels of linoleic (18:2n-6, p=0.001), dihomo--linolenic(p=~.~O5)
and adrenic (p<0.05) acids were reduced. This difference between
the two groups could be due to low intake, impaired accretion
and/or higher turnover in the U C group. Since there was no
difference in the plasma fatty acids between the two groups, diet
may not be the cause. The data indicate that membrane AA and
DHA can be used as biochemical markers to distinguish between
C D and UC.
0 2001 Biochemical Society