Carnitine biosynthesis

Carnitine biosynthesis is a method for the endogenous production of L-carnitine, a molecule that is essential for energy metabolism.[1][2][3][4] In humans and many other animals, L-carnitine is obtained from both diet and by biosynthesis.[5][6] The carnitine biosynthesis pathway is highly conserved among many eukaryotes and some prokaryotes.[7][8][9]

L-Carnitine is biosynthesized from Nε-trimethyllysine.[10] At least four enzymes are involved in the overall biosynthetic pathway. They are Nε-trimethyllysine hydroxylase, 3-hydroxy-Nε-trimethyllysine aldolase, 4-N-trimethylaminobutyraldehyde dehydrogenase and γ-butyrobetaine hydroxylase.

Nε-Trimethyllysine hydroxylase
Main article: Trimethyllysine dioxygenase

The first enzyme of the L-carnitine biosynthetic pathway is Nε-trimethyllysine hydroxylase, an iron and 2-oxoglutarate (2OG)-dependent oxygenase that also requires ascorbate. [11] Nε-trimethyllysine hydroxylase catalyses the hydroxylation reaction of Nε-trimethyllysine to 3-hydroxy-Nε-trimethyllysine.

The current consensus theory about the origin of Nε-trimethyllysine in mammals is that mammals utilise lysosomal or proteasomal degradation of proteins containing Nε-trimethyllysine residues as starting point for carnitine biosynthesis.[12][13][14] An alternative theory involving endogenous non-peptidyl biosynthesis was also proposed, based on evidence gathered from a study involving feeding normal and undernourished human subjects with the amino acid lysine.[15] Although Nε-trimethyllysine biosynthetic pathway involving Nε-trimethyllysine methyltransferase has been fully characterised in fungi including Neurospora crassa, such biosynthetic pathway has never been properly characterised in mammals or humans.[16] A third theory about the origin of Nε-trimethyllysine in mammals does not involve biosynthesis at all, but involves direct dietary intake from vegetable foods. High-performance liquid chromatography (HPLC) analysis has confirmed that vegetables contains a significant amount of Nε-trimethyllysine.[17]

3-Hydroxy-Nε-trimethyllysine aldolase

The second step of L-carnitine biosynthesis requires the 3-hydroxy-Nε-trimethyllysine aldolase enzyme. 3-hydroxy-Nε-trimethyllysine aldolase is a pyridoxal phosphate dependent aldolase, and it catalyses the cleavage of 3-hydroxy-Nε-trimethyllysine into 4-N-trimethylaminobutyraldehyde and glycine.

The true identity of 3-hydroxy-Nε-trimethyllysine aldolase is elusive and the mammalian gene encoding 3-hydroxy-Nε-trimethyllysine aldolase has not been identified. 3-hydroxy-Nε-trimethyllysine aldolase activity has been demonstrated in both L-threonine aldolase and serine hydroxymethyltransferase,[18][19] although whether this is the main catalytic activity of these enzymes remains to be established.

4-N-Trimethylaminobutyraldehyde dehydrogenase
Main article: 4-trimethylammoniobutyraldehyde dehydrogenase

The third enzyme of L-carnitine biosynthesis is 4-N-trimethylaminobutyraldehyde dehydrogenase.[20] 4-N-trimethylaminobutyraldehyde dehydrogenase is a NAD+ dependent enzyme. 4-N-trimethylaminobutyraldehyde dehydrogenase catalyses the dehydrogenation of 4-N-trimethylaminobutyraldehyde into gamma-butyrobetaine.

Unlike 3-hydroxy-Nε-trimethyllysine aldolase, 4-N-trimethylaminobutyraldehyde dehydrogenase has been identified and purified from many sources including rat[21] and Pseudomonas.[22] However, the human 4-N-trimethylaminobutyraldehyde dehydrogenase has so far not been identified. There is considerable sequence similarity between rat 4-N-trimethylaminobutyraldehyde dehydrogenase and human aldehyde dehydrogenase 9,[23] but the true identity of 4-N-trimethylaminobutyraldehyde dehydrogenase remains to be established.

γ-Butyrobetaine hydroxylase
Main article: gamma-butyrobetaine dioxygenase

The final step of L-carnitine biosynthesis is γ-butyrobetaine hydroxylase, a zinc binding enzyme[24][25][26][27][28][29]. Like Nε-trimethyllysine hydroxylase, γ-butyrobetaine hydroxylase is a 2-oxoglutarate and iron(II)-dependent oxygenase. γ-Butyrobetaine hydroxylase catalyses the stereospecific hydroxylation of γ-butyrobetaine to L-carnitine.

γ-Butyrobetaine hydroxylase is the most studied enzyme among the four enzymes in the biosynthetic pathway. It has been purified from many sources, such as Pseudomonas,[30] rat,[31][32][33] cow,[34] guinea pig[35] and human.[36] Recombinant human γ-butyrobetaine hydroxylase has also been produced by Escherichia coli[27] and baculoviruses[26] systems.

Scheme describing the biosynthetic pathway of L-carnitine in humans.
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