Janice Young

Structure determination of a human lymphocyte derived neutrophil activating peptide (LYNAP)

Phytohemagglutinin or Concanavalin A-stimulated human T-lymphocytes produce a factor (LYNAP) with potent chemotactic and enzyme degranulating activity in peripheral human neutrophils. Sequence analysis of LYNAP established an apparently novel 72 residue polypeptide structure. Examination of protein data bases showed that LYNAP had about 30% sequence homology with recently characterised connective tissue activating proteins produced by platelets. Furthermore, it was subsequently found that the amino acid sequence is largely the same as that predicted from a cDNA clone derived from mRNA elevated in peripheral human leukocytes stimulated by mitogens.

Isolation and Characterization of the Intact Gastrin Precursor From a Gastrinoma

Antibodies to the extreme C‐terminal region of human progastrin have been used to monitor the isolation of high‐M r immunoreactive material in a gastrinoma extract. Microsequence analysis of the product revealed amino acid residues in the first 18 positions corresponding to those predicted from the cDNA sequence for preprogastrin starting at position 22; the sequence and immunochemical data together allow the identification of this material as intact progastrin. Implications for gastrin biosynthesis are discussed.

Isolation, structure and properties of the C-terminal flanking peptide of preprocholecystokinin from rat brain

The C‐terminal flanking peptide of preprocholecystokinin has been isolated from rat brain. Micro‐sequence analysis revealed the primary structure: Ser‐Ala‐Glu‐Asp‐Tyr‐Glu‐Tyr‐Pro‐Ser. Arylsulphatase and mild acid hydrolysis suggested that both tyrosine residues are sulphated. The peptide was not active in bioassay systems that respond to CCK8; the significance of the conserved tripeptide Ser‐Ala‐Glu is discussed.

The constitution and properties of phosphorylated and unphosphorylated C-terminal fragments of progastrin from dog and ferret antrum

Antibodies to the extreme C-terminal tryptic (nona-) peptide fragment of porcine progastrin have been used in radioimmunoassay to identify progastrin fragments in dog, ferret and pig antral mucosa extracts and to monitor their purification. In addition to previously characterised phosphorylated and unphosphorylated C-terminal tryptic peptides of porcine progastrin a minor form corresponding to the C-terminal octapeptide (i.e. des-Ser C-terminal nonapeptide) was isolated and characterised. The latter form together with phosphorylated and unphosphorylated forms of the nonapeptides were also isolated and chemically characterised from dog antrum, and the unphosphorylated nonapeptide was characterised from ferret antrum. The primary amino acid sequences of the dog, ferret and pig nonapeptides were identical. In ferret the unphosphorylated nonapeptide predominated, and in dog the phosphorylated form predominated; in pig both forms of the nonapeptide were well represented. Intact progastrin was identified in gel filtration eluates of extracts of all 3 species, but occurred only in relatively low concentrations. The nonapeptides did not stimulate acid secretion in the conscious gastric fistula rat and they did not modify the acid response to G17. Phosphorylation of progastrin-derived peptides is evidently well conserved across a range of species even though there appear to be differences in the relative proportions of phosphorylated and unphosphorylated forms.

Proenkephalin A Processing in the Upper Digestive Tract: Isolation and Characterisation of Phosphorylated N-Terminally Extended Met-Enkephalin Arg6Phe7 Variants

Abstract: Previous studies suggest the processing of proenkephalin A in the porcine upper digestive tract might differ from that in the brain. To characterise more precisely some of the products, we have used antibodies to Met‐enkephalin Arg6Phe7 (MERF) in radioimmunoassay to monitor the isolation of immunoreactive peptides from extracts of porcine pyloric antral muscle, antral mucosa, and duodenum. Seph‐adex G50 gel filtration of each extract produced a single broad peak of high‐molecular‐weight MERF‐immunoreactivity. On anion‐exchange chromatography the antral muscle MERF‐immunoreactivity fractionated into two major peaks, and that from the antral mucosa and duodenum each into four major peaks, suggesting tissue specific processing of proenkephalin A within the porcine gut. Reverse‐phase HPLC and Edman degradation analysis revealed that the least acidic antral muscle peptide was a 31‐residue N‐terminally extended form of MERF that is equivalent to proenkephalin A 209–239. Alkaline phosphatase digestion of the N‐terminally extended MERF variants indicated that some of these peptides were modified by phosphorylation. We conclude that there are complex patterns of proenkephalin A processing in the porcine gut, which in part are due to phosphorylation.

Characterisation of N-terminally extended met-enkephalin Arg6Gly7Leu8 variants in the porcine upper digestive tract

Proenkephalin A-derived peptides are known to occur in the gut, but their precise identity is uncertain. We report here the isolation of N-terminally extended forms of Met-enkephalin Arg6Gly7Leu8 from porcine upper digestive tract monitored by radioimmunoassay. A single major form was identified in pyloric antral muscle and mucosa, but in the duodenum two major forms were detected. Microsequence analysis together with immunological data revealed that the antral mucosal peptide and the most acidic duodenal peptide had identical amino-acid sequences, corresponding to a 5.3 kDa peptide terminating in Met-enkephalin Arg6Gly7Leu8. The data indicate that high-molecular-weight peptides may constitute a major proportion of gut opioid peptide immunoactivity.