Metabolism
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Do we need to synthesise fatty acids?
The simple answer is yes.
There are two enzymes involved in fatty acid synthesis (one is a large multi-functional protein) and there is only one case report of a genetic defect; a newborn girl with severe nerve and muscle damage. Experimental deletion of the gene for one of these enzymes in mice (knock-out mice) is lethal - the knockout embryos are undeveloped at embryonic day 7.5 and die by embryonic day 8.5.
The more interesting question to ponder is the extent to which we need to synthesise fatty acids to maintain adipose tissue stores of triacylglycerol when our diet provides 30 - 45% of energy from fat. Incubation of isolated liver cells (hepatocytes) or adipose tissue cells (adipocytes) with [14C]glucose in the presence of insulin shows that label is incorporated into fatty acids (and then triacylglycerol) in both tissues. In the liver (but not in adipose tissue) label from glucose is also incorporated into glycogen. Therefore, it is obvious that fatty acids can indeed be synthesised from glucose. Similar studies show that fatty acids can be synthesised from acetyl CoA, which is formed in mitochondria from pyruvate, the end-product of glycolysis (see the exercise on life threatening acidosis in an alcoholic).
Adipocytes were incubated with [14C]acetyl CoA and insulin in a conventional phosphate-bicarbonate buffer or a phosphate buffer with no bicarbonate. After 30 min the lipids were extracted into chloroform-methanol and radioactivity was measured in both the lipid fraction and also the aqueous fraction (this will be unmetabolised acetyl CoA). The results were as follows (figures show dpm ± standard deviation for 5 x replicate experiments):
unincubated control (bicarbonate buffer) |
bicarbonate buffer |
bicarbonate-free buffer |
|
aqueous phase |
10,105 ± 120 |
1025 ± 75 |
10096 ± 432 |
lipid phase |
32 ± 16 |
9067 ± 135 |
24 ± 15 |
What conclusions can you draw from these results?
It is obvious that bicarbonate (or carbon dioxide) is essential for fatty acid synthesis from acetyl CoA.
In the next set of experiments adipocytes were incubated with [14C]bicarbonate in the buffer and non-radioactive acetyl CoA, again together with insulin. The results were as follows (figures show dpm ± standard deviation for 5 x replicate experiments):
unincubated control |
30 min incubation |
|
| aqueous phase | 10,295 ± 154 |
10,196 ± 162 |
| lipid phase | 29 ± 15 |
32 ± 14 |
What conclusions can you draw from these results?
Although bicarbonate is required for fatty acid synthesis from acetyl CoA, no label from bicarbonate is incorporated into the fatty acids formed. This suggests that there is an intermediate that is carboxylated, but then loses the carbon dioxide again. The overall reaction can be drawn as:
Two enzymes that are required for fatty acid synthesis were isolated from adipocytes. When both together were incubated with [14C]acetyl CoA in a bicarbonate buffer no radioactive palmitate (C16:0) was formed unless NADPH was added to the incubation mixture. Further studies showed that 14 mol of NADPH were oxidised per mol of palmitate formed.