Generation of Met-enkephalin Arg6Phe7 immunoreactivity by proteolytic cleavage of mammalian plasma precursors by pepsin.
A region-specific antiserum raised against the C-terminal heptapeptide of proenkephalin A (Met-enk Arg6Phe7) was used in RIA studies to show that rat, human, and ovine plasma contain substrates (mol wt, 68K) that yield nanomolar amounts of Met-enk Arg6Phe7 (ME-RF) after treatment with pepsin under acid conditions. This ovine plasma-derived immunoreactivity diluted in parallel to the ME-RF standard in RIA and chromatographed as two low mol wt species (approximately 1K) which were less hydrophobic than the standard on size exclusion and reverse phase chromatography. The pepsin-generated material displaced [3H]naloxone from rat brain binding sites; its potency was about 1000-fold that of ME-RF, assuming near 100% cross-reactivity with the antiserum. Taken together these observations suggest that the pepsin-generated material is of similar mol wt and amino acid sequence to ME-RF, but differs with respect to opiate-binding efficacy, and that the plasma precursor is distinct from proenkephalin in both size and processing sites.
Giraud
A S
AS
Medical Research Centre, Prince Henry's Hospital, Melbourne, Australia.
Parker
L
L
Reichman
C
C
Familari
M
M
Smith
A I
AI
Funder
J
J
D000818:Animals; D002850:Chromatography, Gel; D004744:Enkephalin, Methionine / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology / Q000378:metabolism; D005260:Female; D006801:Humans; D008970:Molecular Weight; D010434:Pepsin A / Q000494:pharmacology*; D011498:Protein Precursors / Q000097:blood*; D011863:Radioimmunoassay; D051381:Rats; D011957:Receptors, Opioid / Q000378:metabolism; D012756:Sheep
D011498:Protein Precursors; D011957:Receptors, Opioid; D004744:Enkephalin, Methionine; C027511:enkephalin-Met, Arg(6)-Phe(7)-; D010434:Pepsin A