Degradation of enkephalins by rat lymphocyte and purified rat natural killer cell surface aminopeptidases.
Enkephalins have been reported to induce an elevation in natural killer (NK) cell cytolytic function. In the central nervous system, the short-lasting biological activity of the enkephalins may be attributable to their rapid hydrolysis at the cell surface. In a similar manner, a potential mechanism for regulating the effects of enkephalins on NK cells is through their degradation at the cell surface. The purpose of this study was to determine if NK cells were capable of enzymatically degrading Met- and Leu-enkephalin and to determine the type(s) of enzymes responsible. We report that rat nylon wool enriched splenocytes, purified NK cells, and interleukin-2 activated NK (A-NK) cells were capable of hydrolyzing the N-terminal Tyr residue from Met- and Leu-enkephalin as determined by high-performance liquid chromatography. The rate of Tyr cleavage from Met-enkephalin was approximately twice that of Leu-enkephalin for both splenocytes, NK, and A-NK cells. On a cellular basis, enkephalin degradation was four to five times greater with A-NK cells than with splenocytes. Only a Tyr cleavage product was detected, which suggested the possibility of an aminopeptidase activity. This was confirmed by the ability of bestatin, a specific inhibitor of cell surface aminopeptidases, to almost completely inhibit enkephalin degradation by splenocytes (85%) and A-NK cells (96%). A-NK cells were more sensitive to bestatin inhibition as indicated by their IC50 values (0.01 mM for splenocytes and 0.001 mM for A-NK cells). In addition, the chelator 1,10-phenanthroline was also capable of effectively inhibiting enkephalin degradation, suggesting that the enzyme responsible has the characteristics of a metalloprotease. In contrast to the effects of bestatin or phenanthroline, typical inhibitors of serine and thiol proteases were without effect.
Amoscato
A A
AA
Pittsburgh Cancer Institute, Pennsylvania.
Spiess
R R
RR
Sansoni
S B
SB
Herberman
R B
RB
Chambers
W H
WH
D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D004745:Enkephalins / Q000378:metabolism*; D007694:Killer Cells, Natural / Q000201:enzymology*; D007700:Kinetics; D008214:Lymphocytes / Q000201:enzymology*; D008297:Male; D008969:Molecular Sequence Data; D015213:Neuroimmunomodulation / Q000502:physiology*; D010446:Peptide Fragments / Q000032:analysis; D051381:Rats; D011916:Rats, Inbred F344 / Q000276:immunology; D013379:Substrate Specificity
D004745:Enkephalins; D010446:Peptide Fragments; D000626:Aminopeptidases