Protein absorption in the intestine is mediated by proteases and brush-border peptidases together with peptide and amino acid transporters. Neutral amino acids are generated by a variety of aminopeptidases and carboxypeptidases and are subsequently taken up by the amino acid transporter B(0)AT1 (SLC6A19), which is mutated in Hartnup disorder. Coexpression of B(0)AT1 together with the brush-border carboxypeptidase angiotensin-converting enzyme 2 (ACE2) in Xenopus laevis oocytes led to a dramatic increase of transporter expression at the oocyte surface. Other members of the SLC6 family were not stimulated by coexpression with ACE2. Addition of a peptide containing a carboxyterminal leucine residue to ACE2- and B(0)AT1-coexpressing oocytes caused inward currents due to Na(+)-leucine cotransport, demonstrating the formation of a metabolic complex. Coexpression of the Hartnup disorder causing mutation B(0)AT1(R240Q) showed reduced interaction with ACE2 and its renal paralogue collectrin. This would result in reduced surface expression in both kidney and intestine, thereby explaining the onset of the disorder in individuals carrying this mutation.

Resorption of amino acids in kidney and intestine is mediated by transporters, which prefer groups of amino acids with similar physico-chemical properties. It is generally assumed that most neutral amino acids are transported across the apical membrane of epithelial cells by system B 0 . Here we have characterized a novel member of the Na ؉ -dependent neurotransmitter transporter family (B 0 AT1) isolated from mouse kidney, which shows all properties of system B 0 . Flux experiments showed that the transporter is Na ؉ -dependent, electrogenic, and actively transports most neutral amino acids but not anionic or cationic amino acids. Superfusion of mB 0 AT1-expressing oocytes with neutral amino acids generated inward currents, which were proportional to the fluxes observed with labeled amino acids. In situ hybridization showed strong expression in intestinal microvilli and in the proximal tubule of the kidney. Expression of mouse B 0 AT1 was restricted to kidney, intestine, and skin. It is generally assumed that mutations of the system B 0 transporter underlie autosomal recessive Hartnup disorder. In support of this notion mB 0 AT1 is located on mouse chromosome 13 in a region syntenic to human chromosome 5p15, the locus of Hartnup disorder. Thus, the human homologue of this transporter is an excellent functional and positional candidate for Hartnup disorder.

The mechanism of the mouse (m)B 0 AT1 (slc6a19) transporter was studied in detail using two electrode voltage-clamp techniques and tracer studies in the Xenopus oocyte expression system. All neutral amino acids induced inward currents at physiological potentials, but large neutral non-aromatic amino acids were the preferred substrates of mB 0 AT1. Substrates were transported with K 0.5 values ranging from approx. 1 mM to approx. 10 mM. The transporter mediates Na + -amino acid co-transport with a stoichiometry of 1:1. No other ions were involved in the transport mechanism. An increase in the extracellular Na + concentration reduced the K 0.5 for leucine, and vice versa. Moreover, the K 0.5 values and V max values of both substrates varied with the membrane potential. As a result, K 0.5 and V max values are a complex function of the concentration of substrate and co-substrate and the membrane potential. A model is presented assuming random binding order and a positive charge associated with the ternary [Na + -substratetransporter] complex, which is consistent with the experimental data.

Transporters of the SLC6 (solute carrier 6) family play an important role in the removal of neurotransmitters in brain tissue and in amino acid transport in epithelial cells. In the present study, we demonstrate that mouse v7-3 (slc6a15) encodes a transporter for neutral amino acids. The transporter is functionally and sequence related to B(0)AT1 (slc6a19) and was hence named B(0)AT2. Leucine, isoleucine, valine, proline and methionine were recognized by the transporter, with values of K(0.5) (half-saturation constant) ranging from 40 to 200 microM. Alanine, glutamine and phenylalanine were low-affinity substrates of the transporter, with K(0.5) values in the millimolar range. Transport of neutral amino acids via B(0)AT2 was Na+-dependent, Cl--independent and electrogenic. Superfusion of mouse B(0)AT2-expressing oocytes with amino acid substrates generated robust inward currents. Na+-activation kinetics of proline transport and uptake under voltage clamp suggested a 1:1 Na+/amino acid co-transport stoichiometry. Susbtrate and co-substrate influenced each other's K(0.5) values, suggesting that they share the same binding site. A mouse B(0)AT2-like transport activity was detected in synaptosomes and cultured neurons. A potential role of B(0)AT2 in transporting neurotransmitter precursors and neuromodulators is proposed.

Neurotransmitter transporters of the SLC6 family play an important role in the removal of neurotransmitters in brain tissue and in amino acid transport in epithelial cells. Here we demonstrate that the mouse homologue of slc6a20 has all properties of the long-sought IMINO system. The mouse has two homologues corresponding to the single human SLC6A20 gene: these have been named XT3 and XT3s1. Expression of mouse XT3s1, but not XT3, in Xenopus laevis oocytes induced an electrogenic Na+-and-Cl--dependent transporter for proline, hydroxyproline, betaine, N-methylaminoisobutyric acid and pipecolic acid. Expression of XT3s1 was found in brain, kidney, small intestine, thymus, spleen and lung, whereas XT3 prevailed in kidney and lung. Accordingly we suggest that the two homologues be termed 'XT3s1 IMINO(B)' and 'XT3 IMINO(K)' to indicate the tissue expression of the two genes.