Chirality differences in amino acid retention and release from acid-extractable pool of cultured mammalian cells.
In previous work, no chiral differences were found between D and L enantiomers of Leu in their ability to displace one another from the acid-extractable pool in mammalian cells. Recent evidence suggested otherwise. Our aim is to examine whether, in physiological range, D-amino acids have an equivalent ability to displace L-amino acids from the acid-extractable pool of HeLa cells, and vice versa. In the Millimolar range, D-Leu and L-Leu have similar uptake and displacement properties with regard to the acid-extractable pool in HeLa cells, despite only the latter isomer being incorporated into protein. Below millimolar concentration 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 at 10(-4) or 10(-5) M displaced and equivalent amount of label from the pool ad D-Leu introduced at a concentration approx. one order of magnitude higher, respectively. Reciprocal experiments, in which the acid-extractable pool was preloaded with 3H-D-Leu, confirmed this finding. The chirality difference was noted whether pool prelabelling was carried out at 37 or 0 degrees C; but in order to avoid the complications of 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, preferential displacement by the L-Leu racemer compared with the D-Leu racemer below mM levels. This was also true for the D and L forms of the non-utilisable isomer of Leu, norleucine (nLeu). We conclude that D-forms of hydrophobic amino acids have lower affinity for similar or the same intracellular binding sites involved in the acid-extractable pool than in their L-forms. The significance of these findings to amino acid pools in cells, and to the predominance of L-forms of amino acids in the biosphere is considered
Wheatley
D N
DN
Cell Pathology Unit, University Medical School, Aberdeen, U.K.
Slater
J
J
Love
E M
EM
Miseta
A
A
D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism* / Q000493:pharmacokinetics; D001693:Biological Transport, Active; D006367:HeLa Cells; D006801:Humans; D007532:Isoleucine / Q000737:chemistry / Q000378:metabolism / Q000493:pharmacokinetics; D007700:Kinetics; D007930:Leucine / Q000737:chemistry / Q000378:metabolism; D009646:Norleucine / Q000737:chemistry / Q000378:metabolism / Q000493:pharmacokinetics; D014176:Protein Biosynthesis; D013237:Stereoisomerism
D000596:Amino Acids; D007532:Isoleucine; D009646:Norleucine; D007930:Leucine