Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01178 (oxytocin)
 15,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A chemical method has been established for the detection of carboxyl-terminally amidated peptides in tissue extracts. Tissue was homogenized in an acidic medium designed to solubilize peptides while precipitating high-molecular-weight protein. The homogenate supernatant was in turn subjected to reversed-phase extraction with C18 Sep-Pak cartridges. The eluates were fractionated by reversed-phase high-performance liquid chromatography (RP-HPLC). Individual fractions were exhaustively digested with thermolysin, derivatized with phenylisothiocyanate (PITC), and then subjected to ethyl acetate extraction under basic conditions. The phenylthiocarbamyl (PTC)-amino acid amide derivatives were selectively taken up into the organic phase, while the other digestion products remained in the aqueous phase. The organic phase was analyzed by RP-HPLC on a Pico-Tag amino acid analysis column, monitoring eluates at 254 nm. PTC-amino acid amides were identified and quantitated by comparing their elution positions and peak areas, respectively, with those of standards. Their identities were confirmed by amino acid analysis, following hydrolysis with hydriodic acid. The technique was applied to extracts of bovine posterior pituitaries and a human medullary thyroid carcinoma. Vasopressin (-Leu-Gly-amide), oxytocin (-Gly-amide), Lys1 gamma 1-melanotropin (-Phe-amide), and various acetylated and non-acetylated forms of alpha-melanotropin (-Val-amide) were identified in the posterior pituitary extract. Various forms of calcitonin (-Val-Gly-Ala-Pro-amide) were detected in the tumour extract. For vasopressin and calcitonin the thermolytic digest resulted in di- and tetra-peptides, respectively, reflecting thermolytic cleavage at more favoured sites.
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PMID:Use of Pico-Tag methodology in the chemical analysis of peptides with carboxyl-terminal amides. 373 29

Bovine neurophysin II was partially digested by chymotrypsin and by chymotrypsin followed by carboxy-peptidase B to produce large fragments collectively representing deletions of residues 1-5 and 91-95. All such fragments were capable of binding peptides to the principal hormone-binding site of neurophysin with normal or near-normal affinity, indicating that residues 1-5 and 91-95 do not directly participate in binding. In addition, preliminary results with thermolysin-derived fragments suggested that residue 6 does not participate in peptide binding. During the course of chymotrypsin studies, it was demonstrated that bovine neurophysin II behaves as a transient competitive inhibitor of chymotrypsin; for neurophysin-peptide complexes, Ki congruent to 8 x 10(-6) M. This inhibition is dependent on neurophysin conformation and is relieved by the anomalous preferential splitting by chymotrypsin of Arg-Arg and Phe-Pro bonds near the carboxyl terminus of neurophysin II. It is suggested that this phenomenon might reflect the interaction of neurophysin II with a chymotrypsin-related enzyme in the pituitary. One approach used in the study of binding properties of proteolytically modified neurophysin was affinity chromatography; the preparation and properties of a conveniently prepared affinity column for neurophysin are described.
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PMID:Partial digestion of neurophysins with proteolytic enzymes: unusual interactions between bovine neurophysin II and chymotrypsin. 707 53