Isolation and characterization of growth hormone from a marine fish, bonito (Katsuwonus pelamis)

Abstract

Growth hormone (GH) was extracted under alkaline conditions (pH 10) from pituitary glands (6.3 g) of bonito (Katsuwonus pelamis), and subsequently purified by gel filtration, ion exchange chromatography, and reversed‐phase HPLC. The GH was monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) and by immunoblotting with yellowtail GH antiserum at each step of purification. GH activity was determined by an in vivo bioassay. The yield of this hormone was 4.8mg/g wet tissue. Intraperitoneal injection of bonito GH at doses of 0.1 and 1 μg/g body wt at 7‐day intervals resulted in a significant increase in body weight and length of juvenile rainbow trout. Bonito GH antiserum exhibited both species and hormone specificity in radioimmunoassay. However, the bonito GH antiserum as well as yellowtail GH antiserum exhibited hormone specificity but not species specificity in immunoblotting. A molecular weight of 21 000 and an isoelectric point of 7.0 for bonito GH were estimated by SDS‐PAGE and gel electrofocusing, respectively. The complete amino acid sequence of 185 residues was determined by sequencing fragment peptides prepared by chemical and enzymatic cleavages. Sequence comparison of bonito GH with other GHs revealed that there is a significant deletion in the middle of the molecule.