CHIRALITY DIFFERENCES IN AMINO-ACID RETENTION AND RELEASE FROM THE ACID-EXTRACTABLE POOL OF CULTURED-MAMMALIAN-CELLS

In previous work, no chiral differences were found between D and L ena ntiomers of Leu in their ability to displace one another from the acid -extractable pool in mammalian cells. Recent evidence suggested otherw ise. Our aim is to examine whether, in physiological range, D-amino ac ids have an equivalent ability to displace L-amino acids from the acid -extractable pool of HeLa cells, and vice versa. In the millimolar ran ge, D-Leu and L-Leu have similar uptake and displacement properties wi th regard to the acid-extractable pool in HeLa cells, despite only the latter isomer being incorporated into protein. Below millimolar conce ntrations however, a distinct difference was found in the displacement of tritium-labelled L-Leu from the pool by unlabelled D-Leu compared with unlabelled L-Leu. Thus, unlabelled L-Leu in the external medium a t 10(-4) or 10(-5) M displaced an equivalent amount of label from the pool as D-Leu introduced at a concentration approx. one order of magni tude higher, respectively. Reciprocal experiments, in which the acid-e xtractable pool was preloaded with H-3-D-Leu, confirmed this finding. The chirality difference was noted whether pool prelabelling was carri ed out at 37 or 0 degrees C; but in order to avoid the complications o f active transport mechanisms, the competition work reported here was done at 0 degrees C. Similar chirality differences were observed with other hydrophobic amino acids, including His, Ile and Phe, such as, pr eferential displacement by the L-Leu racemer compared with the D-Leu r acemer below mM levels. This was also true for the D and L forms of th e non-utilisable isomer of Leu, norleucine (nLeu). We conclude that D- forms of hydrophobic amino acids have lower affinity for similar or th e same intracellular binding sites involved in the acid-extractable po ol than their L-forms. The significance of these chirality findings to amino acid pools in cells, and to the predominance of L-forms of amin o acids in the biosphere is considered. Copyright (C) 1996 Elsevier Sc ience Ltd