133 5 Establishing a peroxisomal β-oxidation computational kinetic model to understand the effects of amino-acid restriction oxygen into hydrogen peroxide, which is then converted back into molecular oxygen by catalase. The resulting 2-trans-enoyl-CoA is further hydrated, dehydrogenated and thiolysed to give a Cn-2 -acyl-CoA and an acetyl-CoA by the bifunctional enzyme (DBP and LBP) and the peroxisomal 3-ketoacylCoA thiolase 1 (ACAA1) and Sterol Carrier Protein X (SCP2), respectively. The resulting Cn-2-acyl-CoA can be further oxidized in the next β-oxidation cycle, or reversibly converted into an acyl-carnitine by the carnitine acetyltransferase (CRAT) or the carnitine octanoyltransferase (CROT). These carnitine esters can be exported out of the peroxisome (Figure 1). Transport of Acyl-CoAs into the peroxisomes Prior to their processing inside the peroxisomes, fatty acyl-CoAs are imported into the organelle. ATP binding cassette (ABC) transporters are membranebound proteins, found in many organelles, catalyzing the transport of lipids, amino acids, and peptides and driven by the hydrolysis of ATP. Within the superfamily of ATP-binding cassette transporters there is a subfamily known as “subfamily D”. Three of these ABCD transporters have been identified in the peroxisome: adrenoleukodystrophy protein (ALDP or ABCD1), ALDP-related protein (ALDRP or ABCD2), and the 70kDa peroxisomal membrane protein (PMP70 or ABCD3) 20,21. ABCD1 and ABCD2 have overlapping specificities for their substrates21. ABCD1 is responsible for the transport of saturated and unsaturated LCFA-CoAs and VLCFA-CoAs into the peroxisome. While there is a clear overlap in substrate specificity between ABCD1 and ABCD2, there are different substrates to which they exhibit their preferences. ABCD2 has a higher specificity for C22:0CoA and poly-unsaturated fatty acids C22:6-CoA and C24:6-CoA20,22.. PMP70 (ABCD3) is one of the most abundant membrane proteins in peroxisomes and it transports the broadest range of substrates. ABCD3 transports saturated and unsaturated LCFA, such as C10:0-CoA, C12:0-CoA and C16:0-CoA, C18:1-CoA, C18:2-CoA, C20:5-CoA and C22:6-CoA23 as well has phytanoyl and pristanoylCoA and dicarboxylic acids24. Moreover, mice depleted of ABCD3 showed accumulation of C27-bile acids in liver, intestine and bile25. There is a clear lack of kinetic information on the peroxisomal ABCD transporters. To date, we have not found any studies describing the kinetic properties of the import of fatty acids into the peroxisome. Measuring transporter kinetics is a laborious process highly affected by lipid membrane dynamics and difficult isolation procedures.
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