Patterns of prohormone processing. Order revealed by a new procholecystokinin-derived peptide.
An 83-amino acid cholecystokinin peptide with a sulfated tyrosine and an amidated carboxyl terminus (CCK-83) was purified from human intestinal mucosa. The purified peptide was chemically characterized, and its bioactivity was compared to CCK-8. Several post-translational processing steps such as cleavage at basic residues, sulfation, and amidation are necessary to form biologically active cholecystokinin from its nascent prepropeptide. The discovery of CCK-83 gives new insight into the order of preprohormone processing. The processing of prepro-CCK appears to be in the order of: 1) signal peptidase cleavage, 2) tyrosine sulfation, 3) cleavage after a carboxyl-terminal pair of basic residues, 4) carboxypeptidase B-like cleavage of these basic residues, 5) amidation (which results in the formation of CCK-83), and 6) cleavage at monobasic residues by endopeptidases (which results in the smaller molecular forms of cholecystokinin). The characterization of biologically active CCK-83 with a sulfated tyrosine and an amidated carboxyl terminus establishes the site of signal peptidase action and suggests an order of post-translational modifications that give rise to the various molecular forms of cholecystokinin.
California Biotechnology Institute, Mountain View 94043.
G A
GA
Eberlein
V E
VE
Eysselein
M T
MT
Davis
T D
TD
Lee
J E
JE
Shively
D
D
Grandt
W
W
Niebel
R
R
Williams
J
J
Moessner
J
J
Zeeh
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000681:Amylases / Q000378:metabolism; D000818:Animals; D002766:Cholecystokinin / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D010450:Endopeptidases / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007413:Intestinal Mucosa / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D010179:Pancreas / Q000187:drug effects / Q000201:enzymology; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping; D011498:Protein Precursors / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012697:Serine Endopeptidases; D012844:Sincalide / Q000494:pharmacology
D000596:Amino Acids; D008565:Membrane Proteins; D010446:Peptide Fragments; D011498:Protein Precursors; D021382:Protein Sorting Signals; C060249:procholecystokinin; D002766:Cholecystokinin; D000681:Amylases; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase; D012844:Sincalide