Bröer S, Palacín M. The role of amino acid transporters in inherited and acquired diseases

Research School of Biology, Australian National University, Canberra, ACT 0200, Australia.
Biochemical Journal (Impact Factor: 4.4). 06/2011; 436(2):193-211. DOI: 10.1042/BJ20101912
Source: PubMed


Amino acids are essential building blocks of all mammalian cells. In addition to their role in protein synthesis, amino acids play an important role as energy fuels, precursors for a variety of metabolites and as signalling molecules. Disorders associated with the malfunction of amino acid transporters reflect the variety of roles that they fulfil in human physiology. Mutations of brain amino acid transporters affect neuronal excitability. Mutations of renal and intestinal amino acid transporters affect whole-body homoeostasis, resulting in malabsorption and renal problems. Amino acid transporters that are integral parts of metabolic pathways reduce the function of these pathways. Finally, amino acid uptake is essential for cell growth, thereby explaining their role in tumour progression. The present review summarizes the involvement of amino acid transporters in these roles as illustrated by diseases resulting from transporter malfunction.

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Available from: Stefan Bröer, Sep 29, 2015
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    • "Among the most interesting transporters in human physiology and pathology there is the Na + -dependent glutamine/neutral amino acid transporter ASCT2 (SLC1A5) previously known in humans as ATB0. This transport system has been identified in human cell systems even though the kinetic properties and substrate specificity are not fully understood [2] [3] [12] while the rodent isoform, besides being studied in cell systems [13] [14] [15] [16], has been also functionally and kinetically characterized in proteoliposomes [9] [17] [18]. Basic functional and kinetic parameters of the kidney rat protein determined in both experimental models correlated well. "
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    ABSTRACT: The human glutamine/neutral amino acid transporter ASCT2 (hASCT2) was over-expressed in P. pastoris and purified by Ni(2+)-chelating and gel filtration chromatography. The purified protein was reconstituted in liposomes by detergent removal with a batch-wise procedure. Time dependent [(3)H]glutamine/glutamine antiport was measured in proteoliposomes which was active only in the presence of external Na(+). Internal Na(+) slightly stimulated the antiport. Optimal activity was found at pH 7.0. A substantial inhibition of the transport was observed by Cys, Thr, Ser, Ala, Asn and Met (≥ 70%) and by mercurials and methanethiosulfonates (≥ 80%). Heterologous antiport of [(3)H]glutamine with other neutral amino acids was also studied. The transporter showed asymmetric specificity for amino acids: Ala, Cys, Val, Met were only inwardly transported, while Gln, Ser, Asn, and Thr were transported bi-directionally. From kinetic analysis of [(3)H]glutamine/glutamine antiport Km values of 0.097 and 1.8 mM were measured on the external and internal sides of proteoliposomes, respectively. The Km for Na(+) on the external side was 32 mM. The homology structural model of the hASCT2 protein was built using as template the GltPh of P. horikoshii. Cys395 was the only Cys residue externally exposed, thus being the potential target of SH reagents inhibition and, hence, potentially involved in the transport mechanism.
    Biochimica et Biophysica Acta 06/2013; 1828(9). DOI:10.1016/j.bbamem.2013.05.034 · 4.66 Impact Factor
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    • "In the cells of different organs, AAs may then serve as building blocks for the synthesis of structural and functional proteins, may be used for cellular metabolism or function as signalling molecules (Verrey et al. 2009; Wu, 2009). Therefore, AA transfer across plasma membranes via various cooperating AA transporters plays a crucial role for body AA homeostasis , and the defect of transporters leads to several diseases (Verrey et al. 2009; Broer & Palacin, 2011). The best characterized basolateral AA transporters of the small intestine and proximal kidney tubule (re)absorbing epithelia are the abundant Lat2–4F2hc (Slc7a8) and y + Lat1–4F2hc (Slc7a7) that function as obligatory exchangers (antiporters). "
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    ABSTRACT: The uniporter TAT1 (Slc16a10) mediates the facilitated diffusion of aromatic amino acids across basolateral membranes of kidney, small intestine and liver epithelial cells and across the plasma membrane of non-epithelial cells like skeletal myocytes. Its role for body amino acid homeostasis was now investigated using newly generated TAT1 (Slc16a10) defective mice (tat1-/-). These mice grow and reproduce normally, show no gross phenotype and no obvious neurological defect. Histological analysis did not reveal abnormalities and there is no compensatory change in any tested amino acid transporter mRNA. TAT1 null mice display however increased plasma, muscle and kidney aromatic amino acid concentration under both normal and high protein diet, although this concentration remains normal in liver. A major aromatic aminoaciduria and a smaller urinary loss of all substrates additionally transported by L-type amino acid antiporter Lat2-4F2hc (Slc7a8) were revealed under high protein diet. This suggests an epithelial transport defect as also shown by the accumulation of intravenously injected 123I-2-I-L-Phe in kidney and of 3H-L-Phe in ex vivo everted gut sac enterocytes. Taken together, these data indicate that the uniporter TAT1 is required to equilibrate the concentration of aromatic amino acids across specific membranes. For instance, it enables hepatocytes to function as sink that controls the extracellular aromatic amino acid concentration. Additionally, it facilitates the release of aromatic amino acids across the basolateral membrane of small intestine and proximal kidney tubule epithelial cells, thereby allowing the efflux of other neutral amino acids presumably via Lat2-4F2hc.
    The Journal of Physiology 10/2012; 590(24). DOI:10.1113/jphysiol.2012.239574 · 5.04 Impact Factor
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    • "which account for about 3 % of the total ORFs. (ii) Defects of transporter functions have been correlated to several human pathologies of a wide range of severity [1–6]. (iii) Recently it emerged that transporters have high relevance in drug delivery, absorption, and side effect onset representing the first contact point with the organism [7–9]. "
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    ABSTRACT: The OCTN subfamily includes OCTN1, 2, and 3 which are structurally and functionally related. These transporters are involved in maintenance of the carnitine homeostasis, which is essential in mammals for fatty acid β-oxidation, VLDL assembly, post-translational modifications, and other essential functions. Indeed, defects of these transporters lead to severe pathologies. OCTN1 and OCTN2 are expressed in many human tissues, while OCTN3 gene has been identified only in mouse and rat. The transporters mediate transport of carnitine and other substrates with different efficiencies and mechanisms. In order to over express the three proteins, a screening of many combinations of E. coli strains with plasmid constructs has been conducted. Only Rosetta(DE3) or Rosettagami2(DE3) gave significant expression. Higher protein amounts were firstly obtained with pET-41a(+) or pGEX-4T1 carrying fusion protein tags which required additional purification passages. Vectors carrying only a 6His tag, suitable for single passage purification, were preferred even though they lead to lower initial expression levels. Expressions were then increased optimizing several critical parameters. hOCTN1 was obtained with pH6EX3 in RosettaGami2(DE3)pLysS. hOCTN2 and mOCTN3 were obtained using pET-21a(+) in Rosetta(DE3). In particular, hOCTN2 was expressed only after codon bias, substituting the second triplet CGG with AAA (R2K mutant). The best growth conditions for hOCTN1 and mOCTN3 were 28 °C and 6 h of induction, while 4 h of induction for hOCTN2R2K. The proteins collected in the insoluble fraction of cell lysates, solubilized with sarkosyl, were purified by Ni-chelating chromatography. Final yield was 2.0, 3.0, or 3.5 mg/l of cell culture for mOCTN3, hOCTN1, or hOCTN2R2K. The data indicated that, in spite of the close evolutionary relations, several factors play different critical roles in bacterial expression of the three proteins, thus general criteria cannot be underlined. However, the strategy of dealing with related proteins revealed to be finally successful for over expressing all the three subfamily members.
    Molecular Biotechnology 07/2012; 54(2). DOI:10.1007/s12033-012-9586-8 · 1.88 Impact Factor
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