Leucine transport

M. Tomi, M. Mori, M. Tachikawa, K. Katayama, T. Terasaki, and K. Hosoya. L-Type amino acid transporter 1 (LATl)-mediated L-leucine transport at the inner blood-retinal barrier. Invest. Ophthalmol. Vis. Sci. 46 2522-2530... 

Effect of an Ionophore on Active Transport Consider the leucine transporter described in Problem 19. Would Umax and/or Kt change if you added a Na+ ionophore to the assay solution containing Na+ Explain. 

Intestinal Uptake of Leucine You are studying the uptake of L-leucine by epithelial cells of the mouse intestine. Measurements of the rate of uptake of L-leucine and several of its analogs, with and without Na+ in the assay buffer, yield the results given in the table. What can you conclude about the properties and mechanism of the leucine transporter Would you expect L-leucine uptake to be inhibited by ouabain ... 

The alternating access transport model has been used to describe the mechanism by which substrates are transported across the membrane via the monoamine transporters (Forrest et al., 2008). This model postulates that the transporter can exist in at least two conformations. These conformations include an extracellularly facing form that is open to the extracellular environment and can bind substrate and Na" and CF ions (Forrest et al., 2008). An intracellularly facing form allows the release of substrate into the cell and the binding of the countertransported ion to reverse the conformation of the transporter (Forrest et al., 2008). The alternating access model is supported by recent crystal structures of other transporters (Weyand et al., 2008 Faham et al., 2008). Two additional conformations of these transporters have also been described. A closed-closed conformation is predicted that prevents accessibility of substrate and ions from either side of the transporter. This closed-closed conformation was observed in the crystal structure of a leucine transporter... 

Indarte M, Madura JD, Surratt CK (2008) Dopamine transporter comparative molecular modeling and binding site prediction using the LeuT(Aa) leucine transporter as a template. Proteins 70 1033-1046... 

Now that we have high-resolution crystal stmctures of membrane transport proteins, we can begin to understand how ion selectivity is accomplished. In Figure 9.2 the Na" -selective binding sites in the Na -dependent leucine transporter LeuT and the K -selective binding sites in the K" " channel provide a direct comparison of selectivity for Na - and K. ... 

Persell R, Haschemeyer AEV (1980) Effects of temperature on L-leucine transport in toadfish liver in vivo. Biochem Biophys Acta 602 650-660... 

We determined the apparent first-order rate constant for the inactivation of the tyrosine/phenylalanine transport system by several concentrations of PCK. By plotting these data in double-reciprocal form we were able to demonstrate the intermediacy of an enzyme-PCK complex in the inactivation reaction and determine K, values of 194 and 177 itM for PCK inhibition of tyrosine and phenylalanine transport, respectively. In addition, this analysis gives the actual first-order rate constants for the inactivation of tyrosine and phenylalanine transport by the enzyme-PCK complex of 0.016 and 0.012 M", respectively. Given the level of imprecision in the assays using whole-cell systems, these numbers are in agreement. The rate of loss of leucine transport activity is comparable to that for tyrosine and phenylalanine transport activity. 

The amino acid L-tryptophan is the precursor for the synthesis of 5-HT. The synthesis and primary metabolic pathways of 5-HT are shown in Figure 13-5. The initial step in the synthesis of serotonin is the facilitated transport of the amino acid L-tryptophan from blood into brain. The primary source of tryptophan is dietary protein. Other neutral amino acids, such as phenylalanine, leucine and methionine, are transported by the same carrier into the brain. Therefore, the entry of tryptophan into brain is not only related to its concentration in blood but is also a function of its concentration in relation to the concentrations of other neutral amino acids. Consequently, lowering the dietary intake of tryptophan while raising the intake of the amino acids with which it competes for transport into brain lowers the content of 5-HT in brain and changes certain behaviors associated with 5-HT function. This strategy for lowering the brain content of 5-HT has been used clinically to evaluate the importance of brain 5-HT in the mechanism of action of psychotherapeutic drugs. 

Scholze, P, Freissmuth, M., and Sitte, H. H. (2002) Mutations within an intramembrane leucine heptad repeat disrupt oligomer formation of the rat gaba transporter 1. J. Biol. Chem. 277,43682 13690. 

In a muscle at rest, most of the 2-oxo acids produced from transamination of branched chain amino acids are transported to the liver and become subject to oxidation in reactions catalysed by branched-chain 2-oxo acid dehydrogenase complex. During periods of exercise, however, the skeletal muscle itself is able to utilize the oxo-acids by conversion into either acetyl-CoA (leucine and isoleucine) or succinyl-CoA (valine and isoleucine). 

Lennernas s group at Uppsala has performed extensive studies to confirm the validity of this in vivo experimental set-up at assessing the rate and the extent of drug absorption. Recovery of PEG 4000 (a non-absorbable marker) is more than 95%, which indicates that the absorption barrier is intact. In addition, maintenance of functional viability of the mucosa during perfusion has been demonstrated by the rapid transmucosal transport of D-glucose and L-leucine. Estimation of absorption half-lives from the measured Pefr agree well with half-lives derived from oral dose studies in humans (i.e. physiologically realistic half-lives). Human Peff estimates are well correlated with the fraction absorbed in humans, and served as the basis for BCS development, and hence the technique is ultimately the benchmark by which other in situ intestinal perfusion techniques are compared. The model has been extensively used to... 

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