Isolation of ACTH1-39,ACTH1-38 and CLIP from the calf anterior pituitary |
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Authors: | P L Brubaker H P Bennett A C Baird S Solomon |
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Institution: | 1. Department of Biochemistry, McGill University Montreal, Canada H3A 1A1;2. Department of Medicine, McGill University, Montreal, Canada H3A 1A1;3. The Endocrine Laboratory, Royal Victoria Hospital, Montreal, Canada H3A 1A1 |
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Abstract: | Calf anterior pituitaries were defatted and homogenized and peptides were adsorbed from the homogenate supernatant onto octadecylsilyl-silica. After elution, the resulting extract was subjected to gradient elution reversed-phase high pressure liquid chromatography (RP-HPLC) using aqueous acetonitrile containing 0.1% () trifluoroacetic acid (TFA). Radioimmunoassay of column fractions for corticotropin (ACTH) revealed three major areas of immunoreactivity. Each was purified to homogeneity by gradient elution RP-HPLC employing aqueous acetonitrile containing either 0.13% heptafluorobutyric acid () or 0.1% TFA (). Amino acid analysis and exopeptidase and trypsin digestions revealed the three forms of corticotropin to be ACTH1–38, corticotropin-like intermediary lobe peptide, (CLIP, ACTH18–39) and ACTH1–39. 3H-labeled ACTH1–39 did not give rise to either 3H-ACTH1–38 or 3H-CLIP during isolation. |
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Keywords: | RP-HPLC reversed-phase high pressure liquid chromatography ODS-silica octadecylsilyl-silica TFA trifluoracetic acid HFBA hepta-fluorobutyric acid corticotropin CLIP corticotropin-like intermediary lobe peptide |
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