The peroxisomal NAD + carrier of Arabidopsis thaliana transports coenzyme A and its derivatives Gennaro Agrimi & Annamaria Russo & Ciro Leonardo Pierri & Ferdinando Palmieri Received: 16 March 2012 / Accepted: 19 April 2012 / Published online: 4 May 2012 # Springer Science+Business Media, LLC 2012 Abstract The peroxisomal protein PXN encoded by the Ara- bidopsis gene At2g39970 has very recently been found to transport NAD + , NADH, AMP and ADP. In this work we have reinvestigated the substrate specificity and the transport prop- erties of PXN by using a wide range of potential substrates. Heterologous expression in bacteria followed by purification, reconstitution in liposomes, and uptake and efflux experiments revealed that PNX transports coenzyme A (CoA), dephospho- CoA, acetyl-CoA and adenosine 3′,5′-phosphate (PAP), be- sides NAD + , NADH, AMP and ADP. PXN catalyzed fast counter-exchange of substrates and much slower uniport and was strongly inhibited by pyridoxal 5′-phosphate, bathophe- nanthroline and tannic acid. Transport was saturable with a submillimolar affinity for NAD + , CoA and other substrates. The physiological role of PXN is probably to provide the peroxisomes with the essential coenzymes NAD + and CoA. Keywords Arabidopsis thaliana . At2g39970 protein . Coenzyme A transporter . Membrane transport . Mitochondrial carrier . Peroxisome Abbreviations CoA coenzyme A FMN flavine mononucleotide NMN nicotinamide mononucleotide PAP adenosine 3′,5′-diphosphate PXN peroxisomal NAD + carrier Introduction In both plants and mammals several cofactors such as NAD + , FAD and CoA are synthesized outside the peroxisomes and must be imported into the peroxisomal matrix where they are essential for important processes. The Arabidopsis thaliana gene At2g39970 encodes a member of the mitochondrial carrier family that is localized in the peroxisomal membrane (Eubel et al. 2008; Reumann et al. 2009; Bernhardt et al. 2012). For a long time, the At2g39970 protein was thought to transport ATP into peroxisomes (Fukao et al. 2001), an assumption that was ruled out in 2008 (Linka et al. 2008). Recently, by means of uptake studies in liposomes reconstituted with the recombinant protein, Bernhardt et al. (2012) have shown that the At2g39970 protein, named PXN, is a peroxisomal NAD + transporter. The same authors found that, in addition to NAD + , PXN transports AMP, ADP and NADH. Furthermore, consistently with the hypothesis that PXN provides peroxisomal β-oxidation with NAD + , a retention of oil bodies and a delay in the degradation of storage oil-derived fatty acids was observed in Arabidopsis pxn null mutant seedlings (Bernhardt et al. 2012). In addition, another study (Mano et al. 2011) in which the Arabidopsis mutant for At2g39970 (APEM3) was also characterized con- cluded that the At2g39970 protein, called PMP38, plays an important role in peroxisomal proliferation. The closest relative of At2g39970 in humans is SLC25A17. We have recently found that SLC25A17 is a transporter of CoA, FAD and, to a lesser extent, NAD + (Agrimi et al. 2012). In this work we have reinvestigated the substrate specificity of the At2g39970 protein, named PXN according to Bernhardt et al. (2012), using a wide range of potential substrates. PXN G. Agrimi : A. Russo : C. L. Pierri : F. Palmieri (*) Department of Biosciences, Biotechnology and Pharmacological Sciences, University of Bari, Via Orabona 4, 70125 Bari, Italy e-mail: fpalm@farmbiol.uniba.it F. Palmieri CNR Institute of Biomembranes and Bioenergetics, Via Orabona 4, 70125 Bari, Italy J Bioenerg Biomembr (2012) 44:333–340 DOI 10.1007/s10863-012-9445-0