Purification and sequence of rat oxyntomodulin.
Structural information about rat enteroglucagon, intestinal peptides containing the pancreatic glucagon sequence, has been based previously on cDNA, immunologic, and chromatographic data. Our interests in testing the physiological actions of synthetic enteroglucagon peptides in rats required that we identify precisely the forms present in vivo. From knowledge of the proglucagon gene sequence, we synthesized an enteroglucagon C-terminal octapeptide common to both proposed enteroglucagon forms, glicentin and oxyntomodulin, but sharing no sequence overlap with glucagon. We then developed a radioimmunoassay using antibodies raised against the octapeptide that was specific for enteroglucagon peptides without cross-reacting with glucagon. Rat intestine was extracted, and one presumptive enteroglucagon form was purified by following the enteroglucagon C-terminal octapeptide-like immunoreactivity through several HPLC purification steps. Structural characterization of the material by amino acid composition, microsequence, and mass spectral analyses identified the peptide as rat oxyntomodulin. The 37-residue peptide consists of pancreatic glucagon plus the C-terminal extension, Lys-Arg-Asn-Arg-Asn-Asn-Ile-Ala. This now permits synthesis of an unambiguous duplicate of endogenous rat oxyntomodulin for physiological studies.
Collie
N L
NL
Department of Physiology, School of Medicine, University of California, Los Angeles 90024.
Walsh
J H
JH
Wong
H C
HC
Shively
J E
JE
Davis
M T
MT
Lee
T D
TD
Reeve
J R
JR
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005934:Glucagon / Q000737:chemistry / Q000235:genetics; D004763:Glucagon-Like Peptides / Q000737:chemistry* / Q000302:isolation & purification*; D006801:Humans; D007082:Ileum / Q000737:chemistry; D007421:Intestine, Small / Q000737:chemistry*; D007583:Jejunum / Q000737:chemistry; D008297:Male; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000302:isolation & purification; D053772:Oxyntomodulin; D052336:Proglucagon; D011498:Protein Precursors / Q000737:chemistry / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D013552:Swine
D009842:Oligopeptides; D053772:Oxyntomodulin; D011498:Protein Precursors; D052336:Proglucagon; D004763:Glucagon-Like Peptides; D005934:Glucagon