Presence of growth hormone-binding proteins in cattle plasma and milk.
The presence of GH-binding proteins (GHBPs) in the plasma of adult cattle was investigated using Sephadex G-200 filtration, Western ligand blotting and Western blotting. The changes in the concentration of GHBP in the plasma of dairy half-sister heifers during the first year of life as well as the presence of GHBP in milk were also investigated. When analytical chromatography (on a 1.6 x 100 cm column) was performed, five peaks of recombinant bovine GH (rbGH)-associated radioactivity were revealed in cattle plasma; the first peak, which appeared near the void volume, was presumed to represent aggregates, the second (M(r) 290 kDa) and the third peaks (M(r) 75 kDa) corresponded to specific rbGH-GHBP complexes; the last two peaks representing free 125I-labelled rbGH and Na[125I]. Western ligand blotting revealed multiple GHBPs. Three major bands were observed at approximately 190, 58 and 31 kDa; an excess of unlabelled hormone blocked the binding of 125I-labelled rbGH. Minor non-specific binding bands were also detected in cattle plasma with molecular weights between 40 and 136 kDa. One monoclonal antibody (8H7) produced against synthetic peptide (amino acids 54-63 of the extracellular domain of the bovine GH receptor) specifically interacted with 190 and 58 kDa bands while the 31 kDa band was not recognized. Finally, Western ligand blots were performed to evaluate the changes in plasma GHBP during the first year of life in 55 dairy half-sister heifers and to identify GHBP in milk. In plasma, the intensity of the 31 kDa band varied greatly between animals while the other specific bands remained stable.(ABSTRACT TRUNCATED AT 250 WORDS)
Molecular Biology and Animal Physiology Unit, Faculty of Agronomy, Gembloux, Belgium.
A
A
Devolder
R
R
Renaville
M
M
Sneyers
I
I
Callebaut
S
S
Massart
A
A
Goffinet
A
A
Burny
D
D
Portetelle
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001345:Autoradiography; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis* / Q000097:blood* / Q000276:immunology; D002417:Cattle / Q000097:blood / Q000378:metabolism*; D002850:Chromatography, Gel; D005260:Female; D007457:Iodine Radioisotopes; D008024:Ligands; D008892:Milk / Q000737:chemistry*; D011994:Recombinant Proteins / Q000032:analysis / Q000097:blood / Q000276:immunology
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D007457:Iodine Radioisotopes; D008024:Ligands; D011994:Recombinant Proteins; C051815:somatotropin-binding protein