Purification of chinook salmon (Oncorhynchus tshawytscha) GH for receptor study |
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| Authors: | Pierre-Yves Le Bail Geneviève Boulard Bruno Barenton Michel Zygmunt |
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| Institution: | (1) Laboratoire de Physiologie des Poissons, INRA, Campus de Beaulieu, 35042 Rennes Cedex, France;(2) Laboratoire de Physiologie Animale, INRA, 9 place Viala, 34060 Montpellier, France;(3) Station de Pathologie de la Reproduction, INRA, 37380 Nouzilly, France |
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| Abstract: | A method for the purification of chinook Salmon (Oncorhynchus tshawytscha) GH, which retains its biological activity, is described. The biological activity was investigated with an established radioreceptor
assay using liver membranes from pregnant rabbits and bovine GH as standard and labelled hormone. The enrichment of the preparation
was checked with electrophoresis (SDS-PAGE). Extraction and further steps were carried out using low molarity alkaline buffer
(pH 8–10, M = 100 mM). Three chromatography steps were performed (Concanavalin-A sepharose, Bio-gel P60, DEAE). Ion exchange
chromatography was performed under isocratic conditions (using a 50 cm column). Two isoforms (sGH1 and sGH2) were isolated.
The purification yield is 0.7% compared to lyophilized pituitaries. The molecule is homogeneous in SDS-PAGE. Contamination
by prolactin, gonadotrophin and corticotrophin is negligible (< 0.5%). It could be demonstrated that the biological activity
of the preparation is maintained since this preparation stimulates the growth of juvenile trout (Salmo gairdneri) and binds specifically (35%) to trout liver membranes. |
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| Keywords: | fish salmon growth hormone biological activity radioreceptor assay purification techniques |
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